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Flour Strength as Influenced by the 
Addition of Diastatic Ferments 



A THESIS SUBMITTED TO THE FACULTY OF THE GRADUATE 
SCHOOL OF THE UNIVERSITY OF MINNESOTA 

By 

FERDINAND A. COLLATZ, B, S., M. S. 

IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR 
THE DEGREE OF DOCTOR OF PHILOSOPHY 



August, 1922 
Chicago, III. 



Flour Strength as Influenced by the 
Addition of Diastatic Ferments 



A THESIS SUBMITTED TO THE FACULTY OF THE GRADUATE 
SCHOOL OF THE UNIVERSITY OF MINNESOTA 

By 

FERDINAND A. COLLATZ, B. S., M. S. 

IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR 
THE DEGREE OF DOCTOR OF PHILOSOPHY 



August, io2_* 
Chicack), 111. 







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LIBRARY OF CONGRESS ( 

RECEIVED i 



DOCUMENTS DiV.otO. 



OCT 171922 i 



Research Fellowship Plan Under Which Fellows of the American 

Institute of Baking Have Been Enrolled as Graduate 

Students of the University of Minnesota. 

The American Institute of Baking in the fall of 1920 detailed L\vo 
research fellows in the chemistry of baking to work on suitable prob- 
lems as graduate students of the University of Minnesota. These 
fellows were regularly registered in the Graduate School of the Uni- 
versity. They pursued such courses as ordinarily constitute a study 
program of candidates for the doctorate in philosophy, majoring in 
the Division of Agricultural Biochemistry. Research problems were 
selected and outlined in conference with their advisors in this Division, 
actual work on the problems being pursued, l)y special agreement, 
chiefly in the laboratories of the American Institute of Baking. Theses 
based on this research were duly presented in partial fulfillment of 
the degree of Doctor of Philosophy, and accepted by committees of 
the Graduate School of the University. These theses are, by agree- 
ment with the graduate faculty of the Division of Agricultural Bio- 
chemistry, published by their respective authors as bulletins of the 
American Institute of Baking. 



TABLE OF CONTENTS 

Page 

I. Introduction: Definition of Hour .stronL;tli 5 

A. Historical 6 

1. The proteins of wheat flour and their physical rehition to flour 

strength 6 

2. The carbohydrates of wheat flour and their relation to flour 

strength • • ■ • 9 

3. Historical review of the diastatic enzymes 13 

a. The Iodine method for the estimation of diastatic activity ... 15 

b. Copper reduction method for the determination of reducing 

sugars formed by the action of diastase 16 

c. Influence of temperature on diastatic activity 17 

d. Influence of acids, bases and salts on diastatic activity 17 

4. Effects of proteolytic enzymes 18 

5. Hydrogen ion concentration 19 

6. Viscosity 19 

II. Experimental 20 

A. The problem 20 

B. Material 20 

1. Description of materials used in these studies 20 

a. Analysis of wheat flours 21 

b. Analysis of malt flour 21 

c. Analysis of malt extract 21 

C. The methods 22 

1. Determination of diastatic activity 22 

2. Determination of proteolytic activity 23 

3. Determination of gas producing capacity of flour 24 

4. Baking tests 24 

D. Influence of varying conditions on diastatic activity 25 

1. Determination of the optimum pH for the amylase of malt flour.. 25 

2. Influence of time of digestion on the diastatic activity of malt 

flour ; 26 

3. Effect of temperature upon diastatic activity 28 

4. Effect of concentration of diastase on hydrolysis of starch in 

wheat flour 30 

5. Effect of increasing amounts of malt flour when digested with 

a constant quantity of wheat flour 31 

6. Production of reducing sugars in the dough during fermentation .32 

7. Determination of proteolytic activity as measured by the fall in 

viscosity of flour-water suspensions when digested with diastatic 
preparations .41 

8. Gas production capacity of wheat flour in relation to strength.. 47 

9. Change in hydrogen ion concentration of fermenting dough 48 

10. Baking data 4g 

III. Discussion 54 

1. Changes in pH, temperature and concentration and their effects 

upon the activity of diastases contained in malt flour 54 

2. Effect of diastatic enzymes upon starch of different flours 55 

3. Production of reducing sugars in the panary fermentation of bread 

and the effects of diastases added to the dough 56 

4. Eifect of proteolytic enzymes contained in malt preparations 

upon the viscosity of strong and weak flours, following the addi- 
tion of various amounts of lactic acid 58 

5. Gas producing capacities of strong and weak flours and the effect 

of added malt extract upon them 59 

6. Changes in hydrogen ion concentration taking place during the 

fermentation of the dough .60 

7. Effect of malt flour and malt extract upon tlie baking value of flour.60 

IV. Summary ^2 

V. Literature cited 63 



FLOUR STRENGTH AS INFLUENCED BY THE 
ADDITION OF DIASTATIC FERMENTS 

By Ferdinand A. Collatz 

I. INTRODUCTION. 

The baking strength of flour has received a great deal of attention 
by scientific workers in the last twenty-five years, due primarily to 
the economic importance of bread. A number of factors have been 
thoroughly investigated, in their relation to baking strength, in order 
to draw some conclusions as to why some flours give a large, light, pal- 
atable loaf of bread and others an inferior loaf. Certain factors which 
have been investigated in their relation to baking strength are total 
nitrogen, ratio of water-soluble nitrogen to total nitrogen, chemical 
composition of the individual proteins, total gluten, total gliadin, ratio 
of gliadin to glutenin, ratio of gliadin to total nitrogen, ratio of wet 
to dry gluten, efifect of electrolytes, hydrogen-ion concentration, total 
amount of gas evolved during fermentation, and the effects of diastatic 
and proteolytic enzymes of the flour. 

Flours which bake out well have been given the arbitrary term of 
strong flours while the others arc termed weak. Naturally a great 
number of definitions of strength have found their way into the litera- 
ture, but the definition that has been most generally accepted is that of 
Humphries and Biffin (1907), who state that "a strong wheat is one 
which yields flour capable of making large, well-piled loaves." Flours 
which do not measure up to this empirical standard are classed as 
weak. This definition indicates that strength in flour is more desir- 
able than weakness for the baking of bread. Wood (1907), has called 
attention to two factors in strength, namely, size and shape of the loaf. 
This has stimulated a great deal of research by Ford and Guthrie 
(1908), Baker and Hulton (1908), on the diastatic and proteolytic en- 
zymes in wheat flour, and also by Upson and Calvin (1915) (1916), 
Gortner and Doherty (1918), and Sharp and Gortner (1922), on the 
colloidal properties of wheat gluten as affecting flour strength. To- 
day we must recognize three groups of factors dealing with strength 
or weakness in flour, According to Sharp and Gortner (1922). we 
have "at least three classes of weak flour, i. e., (1) weakness due to 
an adequate quantity of gluten but of inferior quality, (2) weakness 
due to an inadequate quantity of a good quality gluten and (3) weak- 
ness due to factors influencing yeast activity." 



HISTORICAL. 

Not until Osborn and \\)urhees (1893) (1894), established the com- 
position and properties of the wheat proteins was any great advance 
made in regard to flour strength, and naturally attention was then di- 
rected to the two main proteins, gliadin and glutenin. Fluerent (1896) 
asserted that flour strength depended upon the proportions of gliadin 
to glutenin, the ratio of 75 percent to 25 percent or 3 to 1 being most 
nearly ideal. Snyder (1899) came to similar conclusions but stated 
that the ideal ratio was 65 [percent gliadin and 35 percent glutenin. 
In a later publication Snyder (1901) claims quality rather than quan- 
tity of protein is of the greater importance. Shutt (1904) (1907) 
points out that after several years of research "it appears extremely 
doubtful if the gliadin number (percentage of albuminoids in the form 
of gliadin) constitutes a factor that can be correlated with bread mak- 
ing values as obtained l)y baking trials." These conclusions are again 
verified in a later report. Thatcher's (1907) results show that no single 
factor or group of chemical tests which he tried give results from 
wdiich the comparative baking qualities (jf flour can be determined. 
Blish (1915) states that the gliadin-glutenin ratio is much more con- 
stant in flours of different baking strength than has heretofore been 
supposed. Blish found, after careful investigation, that the individual 
proteins of weak and strong flours are chemically identical. 

The soluble proteins of flour have also had their share of investiga- 
tions as to their relation to baking strength. Snyder (1897). Bremer 
(1907), Wood (1907) and others have found no relation of baking 
strength to the amounts of water soluble proteins. 

Quite recently Martin (1920) has attemi)ted to correlate certain 
I)roperties of flours with baking strength. He finds "for flours having 
a satisfactory gas producing capacity, bakers' marks, gas retaining ca- 
pacity, and amended gliadin content are closely related, and it is con- 
sidered that the estimation of the gas producing capacity will indicate 
the strength of the flour." 

Martin's "amended gliadin'' content is the dift'erence in protein 
(Nx5.7) between the amounts extracted by 50 percent alcohol and 
that extracted by water acting for three hours at 24-25°. Sharp and 
Gortner (1922) find that "amended gliadin" values were not correlated 
with the strength of the flours with which they worked. 

The Proteins of Wheat Flour and Their Physical Relation to Flour 

Strength. 

As far as we know, all proteins belong to that class of colloids known 
as emulsoids and more recently termed "hydrophylic colloids." As 
the latter term suggests, this class of colloids has a great affinity for 



water, which, however, can be moditied to a great extent by the addi- 
tion of acids, bases and salts to the dispersion medium. 

Hofmeister was one of the first to investigate the swelling of pro- 
teins. He found that in solutions of sulphates, tartrates, acetates, 
alcohol and cane sugar, gelatin-plates take up less water than they do 
when immersed in distilled water, while in solutions of potassium, so- 
dium or ammonium chlorides, sodium chlorate, sodium nitrate and so- 
dium bromide, they take up more water than they do when immersed 
in distilled water. Hofmeister's work has been enlarged upon by oth- 
ers, notably Pauli (1899) (1902) (1903) (1905) (1906) and Fischer 
(1915) (1918). 

Wood (1907) and Wood and Hardy (1908) have demonstrated 
wheat gluten to be an emulsoid colloid and as has already been noted 
the water-holding capacity of hydrophylic colloids can be altered to a 
marked degree by the addition of electrolytes. Acids and bases cause 
imbibition up to a certain point, while neutral salts tend to inhibit the 
imbibition of water. 

It appears that Wood (1907) was the first to call attention to the 
physical properties of the proteins in wheat flours rather than the 
chemical differences in relation to flour strength. He investigated the 
possible chemical differences between the glutcnin and the gliadin of 
these two classes of flours. From this he concludes that strength 
(particularly the shape of the loaf) is closely related to the physical 
state of the gluten, which in turn is affected by the presence of elec- 
trolytes. 

Wood, and Wood and Hardy determined the effects of acids, with 
and without salts, by suspending bits of gluten from glass rods in the 
liquid to be investigated. They found that dispersion of the gluten 
starts immediately when immersed even in the lowest concentration 
of acid, and dispersion increased with increase in concentration within 
certain limits. This holds good for sulphuric, phosphoric and oxalic 
acids, but not for hydrochloric. When the concentration of the latter 
exceeded N/30 the dispersion began to decrease until at a concentra- 
tion of N/12 the gluten was more elastic and coherent than in its 
original condition. The addition of salts decreases dispersion in all 
cases and such amounts of salts can be added which will prevent the 
dispersion of the gluten. From this Wood suggests "that the varia- 
tion in coherence, elasticity and water content, observed in gluten ex- 
tracted from different flours, is due to varying cancentrations of acids 
and salts in the natural surroundings of the gluten, rather than to any 
intrinsic difference in the composition of the glutens themselves." 
Wood thinks that the direct addition of acids or salts to the flt»ur, in 
order to strengthen it, is impractical as they would have to be in con- 



tact for forty-eight hours before baking. lie found that this was the 
time required for the gluten to come into equilibrium with its sur- 
roundings. He, therefore, advocates the blending of such fiours which 
will supply each flour's requirements in this respect. 

In a later paper Wood and Hardy ( 1908) take up the theoretical 
discussion of the efl:'ects of acids, alkalis, and salts up(Mi gluten. 

Upson and Calvin were the next to study the colloidal swelling of 
gluten. They atacked the problem in a slightly different manner, 
washing out the gluten with distilled water and pressing it out into 
thin layers. They next cut out discs of uniform size and weighed 
them immediately. The discs are then immersed in acid and acid-salt 
solutions of varying concentrations for a detinite period of time when 
they are taken out, drained and weighed. They tind that "flours con- 
taining acids and salts in such combinations as to favor water absorp- 
tion will behave as weak flours, wdiercas those containing acids and 
salts in such combinations as inhibit water absorption will behave as 
strong flours." Their conclusions are very similar to those of Wood. 

Gortner and Doherty were the next to investigate the colloidal prop- 
erties of wheat flour gluten. Their method of attack was like that of 
Upson and Calvin, in that they recorded the increase in imbibition by 
weighing discs of gluten after immersion in acid and acid-salt solutions 
of various concentrations. They worked with five different flours, 
namely a high grade |)atent flour milled from hard spring wheat, a 
clear flour and three typically soft flours milled from Oregon wheat. 
Their results show that the gluten from a weak flour has a much low- 
er rate of imbibition and a much lower hydration capacity than the 
gluten from a strong flour; also that inorganic salts when added to an 
acid solution lower the relative imbibition of gluten placed in such 
solutions. Glutens from the different flours react differently to the 
addition of inorganic salts. This leads them to believe "that a weak 
gluten does not owe its 'weakness' nor its imbibition curve its "flat- 
ness,' to either the acid or the salt content of the flour from which it 
is derived, 'but rather to the fact that a weak gluten has inherently in- 
ferior colloidal properties." 

In 1918 and 1919, a series of articles dealing with the physical prop- 
erties of wheat flours were published by Henderson and his co-work- 
ers. Inasmuch as they fail to describe their flours, no conclusions can 
be drawn as to their effect on fl()ur strength. It is of interest to note, 
however, that at a hydrogen ion concentration of about pH 5, (the op- 
timum hydrogen ion concentration in the making of bread as found 
by Jessen-Hanson) the viscosity of dough, made from four different 
flours is at a minimum. They further note that salts such as sodium 
lactate, NaoSO^ MgSC)^, KBrOs tend to decrease the viscosity of the 



dough, while NaCl, MgClz NH^Cl tend to decrease the viscosity 
when used in small amounts, but on further addition the viscosity 
increases. 

Ostwald (1919) and Liiers (1919) and Liiers and Ostwald (1919) 
(1920) in a- series of papers show the remarkable parallelism existing 
between viscosity measurements of flour-water mixtures and grade of 
flour. They found that flours divide themselves into three distinct 
groups when measured in this way. The low extraction flours (40%- 
60%) constitutes one group, the high extraction flours (60%-94%) 
a second, and the tailing flours (remains of 60 % - 94 %) constitute 
the third group. They also find that acids and bases tend to in- 
crease the viscosity of the flour-water mixtures while salts tend 
to depress the viscosity. Their results, although obtained by a dif- 
ferent method, confirm the work of Wood, Wood and Hardy, and Up- 
son and Calvin. It must be emphasized, however, that their results 
were obtained on flours of dift'erent milling extraction and the dif- 
ferences they obtained refer only to the grade or degree of extraction 
of the flours, and do not necessarily apply directly to the problem of 
flour strength. 

Sharp and Gortner have continued the work of Gortner and Do- 
herty, and confirm the findings of the latter in regard to the action of 
acids upon gluten. They find a marked difiference in the rate of im- 
bibition when using the various alkali hydroxides. The action of al- 
kalis on gluten is markedly different from that of acids, as dispersion 
takes place at much lower concentrations. They remark in this re- 
inspect. "Indeed dis]>ersi()n and imbibition arc here almost coincident." 
During the course of the work Sharp and (iortner washed out the 
gluten from their strong and weak flours and dried it at low temper- 
ature in vacuo. After pulverizing, they found that the dried glutens 
of various flours were much more alike than in the wet state. This 
observation is in accord with their theory '"that the strong gluten is 
strong because of its colloidal properties ; inasmuch as it is well known 
that the alternate wetting and dr}ing of a colloidal gel. breaks down 
the gel structure." They also find that the optimum hydrogen ion con- 
centration for imbibition is the same for the different acids used. 

In their next paper Sharp and Gortner (1921) extend their work on 
the hydration capacity of glutens. They find that the viscosimeter 
gives an accurate measure of imbibition in that the curve obtained fol- 
lowed the previous curves by weighing out the discs of gluten. As 
might be expected, they found that the strong flours give much greater 
viscosity measurements than do the weak flours. 

The Carbohydrates of Wheat Flour and Their Relation to Flour 

Strength. 

The carbohydrates in the flour have also been investigated with ref- 

9 



erence to their rule in flour strength. Wood was perhaps the first to 
make any definite statement in this respect, although Girard and Fleu- 
rent (1903) called attention to the great variations in the amounts of 
sugars in the flours. They found on analysis that glucose and cane 
sugar were present, the former varying from 0.097o to 0.81%, the latter 
0.63% to 1.897o. Bruying in 1906 considered that the sugar present in 
flour is not glucose and sucrose, but almost entirely maltose. Liebig 
(1909) supports the views of Girard and Fleurent in that he found 
wheat flour to contain from 1-1.5% sucrose and 0.1-0.4% dextrose. 
Wood thinks that strength hinges to a great extent, as far as volume 
and size of the loaf is concerned, upon the sugar content and the dias- 
tatic enzymes that the flour contains. He sums up that factor in 
strength dealing with volume of the loaf as follows : "The factor which 
primarily determines the size of the loaf which a flour can make is 
quite distinct. The size of the loaf is shown to depend in the first 
instance on the amount of sugar contained in the flour together with 
that formed in vhe dough by diastatic action. Particular attention 
should be paid lo the rate of gas evolution in the later stages of fer- 
mentation, as this is shown to be more directly connected with the size 
of the loaf." Wood's method of measuring the gas-producing capac- 
ity of a flour consists of mixing 20 grams of flour and 0.5 grams of 
yeast with 20 cc of water in stoppered flasks and measuring the lib- 
erated carbon dioxide under brine. 

Shutt (1907) shortly after Wood's article, determined the sugar ex- 
tracted by 70 per cent alcohol and by water and could find no evidence 
"that with increase of sugars there is increase of volume in loaf, but 
rather the reverse." .Shutt's data is shown in Table I. 

TABLE I. 

Sugars in flour extracted by 70 per cent alcohol and water. 
(Data taken from Shutt [1907] page 20). 

In aqiK-ous extract In alcoholic extract 





Directly 


.\fter 




Directly 


After 






Designation 


redncint; 


Inver.sion 




reducing 


Inversion 






of 


as 


as 


Total 


as 


as 


Total 


Vol. 


Sample 


IMaltosc 


Sucrose 


Sugars 


Maltose 


Sucrose 


Sugars 


Bakers 




% 


% 


% 


% 


% 


% 


Mark 


No. 1 Hard 


1 .Q6 


2.14 


4.10 


.13 


.91 


1.04 


492 


No. 1 Northern 


2.73 


1.9.^ 


4.68 


.20 


.94 


1.14 


443 


No. 2 Northern 


1.87 


1.79 


3.66 


.18 


.87 


1.05 


438 


No. 3 Northern 


3.42 


2.10 


.S.22 


.26 


121 


1.47 


383 


No. 4 Com'cial Gi 


-. 3.62 


2.43 


6.05 


.05 


134 


1.39 


397 


No. 5 Com'cial Gi 


-. 3.63 


2.43 


6.06 


.06 


1 42 


1.48 


366 


No. 6 Com'cial Gi 


r. 4.07 


2.43 


6.50 


.06 


1.36 


1.42 


363 



Shortly after the appearance of Wood's paper. Baker and Hulton 
(1908) reported a paper in which they investigated the action of en- 



10 



zynies contained in flour with regard to their etifect on flour strength. 
Unable to demonstrate experimentally the existence of proteolytic en- 
zymes in flour, they concluded that any which are present have no ef- 
fect upon the gluten during the time of fermentation. They did 
show, however, that the proteolytic enzymes contained in yeast play 
an important part, since one dough containing yeast showed 2.7% sol- 
uble nitrogen as protein, while a similar dough without yeast had 
1.9% soluble nitrogen as protein. Although the fact was known that 
wheat flour contained amylolytic enzymes, Baker and Hulton demon- 
strated their presence in dough by an increase in maltose, extracting 
the latter with water and preparing the maltosazone. They found 
that contrary to expectatinns, the diastatic activity of flours increased 
with age. * 

Baker and Hulton (1908) show that the total volume of gas pro- 
duced (same method as outlined by Wood) increases roughly with in- 
crease in baking strength (Baker's Mark) of the flour. They also 
point out that a weak flour may have a diastatic power as high or even 
higher than a strong flour. This is explained by the fact that in real- 
ity the weak flour is deficient in liquifying enzymes and that by the 
addition of liquifying enzymes a much greater volume of gas is given 
off, while a strong flour shows no increase in gas production when a 
diastatic enzyme is added. The liquifying enzymes were added in the 
form of a malt extract and Table II shows that even such smaller 
amounts as 0.25 and 1.0 percent caused the gas production to increase 
enormously in a w^eak flour. They did not state whether the addition 
of the malt extract did actually increase the baking strength of the 
flour. From the data submitted, Baker and Hulton concluded that 
weak flours in some instances give as great a gas production as do 
strong flours, and that gas production is not a function of the quan- 
tity of diastases but, as they show, (Table III) it is intimately con- 
nected with "the additional matter rendered soluble during the process 
of doughing," i. e. maltose. 

TABLE II. 

Effects of added malt extract upon a weak flour with reference to an 

increase in gas production. (From data of Baker 

and Hulton [1908] page 372). 












,25 Percent 


1.0 Per cent 


Time 


FI 


our Alone 




of Malt 


of Malt 


Hours 




c.c. CO. 




c.c. CO2 


c.c. C0= 


0.5 




28 




30 


32 


1.0 




47 




66 


69 


1.5 




55 




101 


115 


14.0 




113 




245 


362 



*(It is a well-known fact that flour.s on aging show greater baking strength 
and this increase in diastatic activity may therefore be the primary cause for 
increase in baking strength a."^ the flour ages. At any rate it seems to be in 
accord with Wood's theory). 

n 



TABLE III. 

The relation between gas volume and the additional matter rendered 

soluble during the process of doughing. (From Baker and Hulton 

(1908) page 372). 









I'ercent 


of 














Matter sc 


)luble 


Differences 


Volume 






Percent 


of 


in dou; 


gli 


Maltose 


of gas ob- 






matter sol 


uble 


when k( 


-Pt 


formed 


tained from 






in water 


at 


at 40^ C. 


for 


durinj.; 


dough in 


Bakers 


our 


15.5°C. 




3 liour.s 


doughing 


3 hours 


Mark 


1 


2.12 




3.60 




1.48 


78 


45 


X 


2.03 




4.41 




1.38 


84 


40 


w 


2.83 




5.38 




2.53 


145 


Id 


3 


2.49 




5.53 




3.04 


155 


80 


Y 


2.69 




6.57 




3.88 


164 


95 


2 


3.19 




6.66 




3.45 


175 


78 


4 


4.19 




10.95 




6.75 


193 


90 


V 


2.83 




8.26 




5.42 


217 


90 


T 


2.84 




7.66 




4.82 


220 


80 


U 


2.65 




7.68 




5.02 


230 


91 


Z 


3.54 




11.65 




8.11 


270 


90 



It would seem from the above table that low strength flours are de- 
ficient in liquifying enzymes and the authors conclude that the liqui- 
fying enzymes are the limiting factors in the production of maltose in 
the dough stage. 

Simultaneously with the appearance of Baker and Hulton's article, 
the work of Ford and Guthrie (1908) was published on the relation 
of enzymes contained in flour to its baking strength. They conducted 
experiments of extraction and found that amylases could be greatly 
stimulated by the use of KCl and also 1)y active and by boiled papain. 
In testing amylase values from twelve flours, they found differences 
(using KCl and papain extracts) varyiii!^ from 22.1 to 4().8 expressed 
in grams maltose per grani of dry flour. They could not correlate 
diastase activity with flour strength and state "It however indicates 
that in developing a method of evaluation, the total amylase is one im- 
l^ortant factor, also that the presence of a proteolytic ferment is an- 
otlier and possibly more valuable consideration." 

Ford and Guthrie (1908) were prol^ably the first to demonstrate 
the action of proteolytic enzymes in flour. They were unable to 
secure results with nitrogen determinations or with the viscosi- 
rneter, so they tried 1 percent gelatin. The liquification of the gela- 
tin gave them positive proof of proteolysis. They also conducted 
baking tests with a large amount of protease added, and naturally 
the loaf did not rise during the fermentation period, the resultant 
bread being a soggy mass. They concluded that proteases decrease 
gas holding properties of the gluten and point out that this is the 
chief reason for failure in the use of malt extracts in baking practise. 

Bailey and Weigley (1922) fotind that flour strength depended 



12 



upon factors which control the rate of carbon dioxide production 
and the amount of carbon dioxide lost during fermentation. They 
found that "the loss of carbon dioxide per unit increase in volume 
under controlled conditions afifords a useful measure of gas-holding 
capacity of dough." 

In some unpublished data Thatcher and Kennedy show that 
when flour was digested with water, the amounts of reducing sugars 
in the extract increased regularly with increase in temperature. A 
centrifuged aliquot of a flour water extract likewise increases in 
soluble nitrogen with increase in temperature of extraction. They 
also found that no increase in reducing sugars takes place when a 
filtered extract (0°-5°C) of flour is allowed to act on soluble starch 
when incubated at 40°, 50° and 60° C. Under these conditions they 
assume that absorption of the enzyme or activator has taken place 
upon the filter paper or uixm the gluten colloids. 

Historical Review of the Study of Diastatic Enzymes. 

In taking up the history of the diastases, one is confronted with 
a voluminous and at times conflicting literature, which extends back 
over a period of one hundred years or more. Naturally, a great 
deal has to be discarded, as it would be impossible to review any but 
the most important papers submitted on this question. Neverthe- 
less it is my intention to cite a number of papers which are of in- 
terest from a purely historical viewpoint. 

Vauquelin in 1811 was the first to record the fact that when 
starch was heated in water, it gave an opaque solution and had the 
characteristics of gum arabic. In the same year Kirchofif found 
that when starch was boiled with dilute H^SQ^j, a crystallized 
sugar was formed. Two years later he noted that the protein of 
the embryo of the seed, particularly if the seed had been germinated, 
acted on starch in much the same manner as did the acid. He real- 
ly was the first to record diastatic activity but did not realize the 
importance of his observations. Vogel in 1812 found that when 
starch was boiled with acid, it gave two products, a sugar and a 
gummy substance, the latter now known as dextrin. Stromeyer 
in 1813 found that iodine was a specific reagent for starch and visa 
versa, while the action of alkaline copper sulphate w^as found by 
Trommer in 1841 to be a means of distinguishing sugar from starches 
and gums. 

The gummy substance found by Vogel was investigated by Biot 
and Persoz in 1853 and was found to turn the plane of polarized 
light to the right. For this reason it was given the name of dex- 
trin. It is of interest to note that the work of Biot and Persoz 
formed the basis for the development of our present day polaris- 

13 



cope. In the same year Payen and Persoz conclusively established 
the fact that an extract of malted grain had a powerful action in 
liquifying and saccharifying starch. They ascribed this function 
to some inner substance and named it diastase. It had been the 
impression of chemists up until this time that glucose was the sugar 
formed when starch was acted upon by diastase and it was not until 
1872-1876 that O'Sullivan showed it to be maltose. O'Sullivan 
found the optical rotation to be too high and the reducing power 
too small to correspond with glucose. It might be of interest to 
call attention to the discovery of maltose at this time. Although 
DeSassure had accurately described maltose in 1819 the fact had 
evidently been forgotten until Dubrunfaut called attention to it in 
1847 and named it maltose. This rediscovery was again forgotten 
until it was again described by O'Sullivan in 1872. 

Marker in 1877 states that at a temperature of 60° C four mole- 
cules of starch yield three of maltose and one of dextrin, under 
the influence of diastatic ferments. At 65" the }'iel<l of maltose is 
lowered and at still higher temperatures two molecules of starch 
yield one of dextrin and one of maltose. Marker concludes that 
there are two diastatic ferments, one producing dextrin and the oth- 
er maltose. This is our present day conception of the diastases, one 
being termed the liquifying and the other the saccharifying enz3'me. 
Musculus and Gruber in 1878 regarded starch as a polysaccharide, 
containing live or six times the group CioHooO,,,. Under the action 
of diastase or acids, the carbohydrate undergoes a series of changes 
of hydration and successive decomposition, resulting in maltose and 
dextrin of less molecular weight. Brown and Heron in 1879 found 
that the heating of a diastatic solution diminishes its activity and 
that an increase in temperature increases its activity up to 66°, beyond 
which not much activity is shown. They also found that alkalies 
markedly reduce the activity <)f a malt extract. 

It was in 1879 that Kjeldahl stated his law of proportionality 
in regard to the action of diastases. He determined the reducing 
power of malt extract and saliva on an excess of starch at 57°-59°. 
He conisdered that the reduced copper was directly proportional to 
the amount of amylase present and was a true measure of diastatic 
power so long as digestion was not carried aliovc 40 i)er cent of the 
starch present, (iriefsmayer a year later, confirmed Kjeldahl's work 
and the law of proportionality. 

Up to about this time the polariscope and the cupric reducing 
method had been used to estimate the diastatic power of malt ex- 
tract by the amount of maltose formed. The iodine reaction was 
made use of by Roberts in 1881, however, and he defined the dias- 

14 



tatic power of pancreatic extracts, saliva, and malt extracts, as the 
number of cubic centimeters of a standard starch paste which could 
be converted by one cubic centimeter of the active solution during 
five minutes at 40° into products giving- no color reaction with 
iodine. 

Jungk two years later published a method of determining the 
diastatic activity of a malt extract by the iodine method which was 
similar to the method of Roberts. He determined the time re- 
quired for 10 cc of extract to convert 10 grams of starch which he 
considered should not exceed 10 minutes, for a good malt. His 
temperature of digestion was 40°. 

From the literature already cited, two methods of determining 
the diastatic activity of an amylase preparation had come into use, 
namely the iodine or the so-called liquifaction method, wdiich mea- 
sures the power of the amylase to completely convert the starch into 
products which no longer give the characteristic color with iodine, 
and the reduction or saccharification method in which the amounts 
of reducing sugars are estimated b}' means of alkaline copper 
sulphate. The development of the iodine method from the time of 
Jungk will be the first considered. 

Iodine Method for the Estimation of Diastatic Activity. 

Francis in 1898 made the next improvement in the iodine method 
by laying down very exact rules for the determination of the end 
point in the iodine starcli reaction. He also extended the time of 
digestion from 10 minutes to half an hour. Takamine in the same 
year developed a different procedure. He first standardized a sample 
of taka-diastase, which was found to keep its diastatic power for a 
considerable length of time. He then determined the relative amounts 
of standard and sample to be tested, which are required to accomplish 
the same amount of conversion in the same length of time. In these 
determinations he used iodine to test for the end point. 

The next important work was carried out by Wohlgemuth in 1908. 
He established a new standard which was based on the number 
of cubic centimeters of 1 percent starch solution which 1 cc of dias- 
tatic ferment could convert in 30 minutes at 40° C. The method 
consists of measuring out 5 cc of a 1 percent starch solution into 
each of a series of test tubes and then adding different amounts of 
a diastatic solution to each. At the end of a half hour at 40° the test 
tubes are transferred to an ice bath. After cooling each tube was 
shaken up with a definite amount of iodine solution and the one 
which showed no trace of color was taken for the end point. 

Johnson in 1908 improved the technique of Francis, and Jungk, by 

15 



preparing a starch paste (potato) of constant value. He added his 
diastatic preparations to a fixed amount of starch and withdrew 
portions and tested with iodine, at the end of ten minutes. When 
close to the end point, he tested more frequently, repeating with 
smaller increments of sample, until the amount was found which 
would in ten minutes just convert the starch. 

Sherman and Schlesinger (1913) and Sherman and Thomas (1915), 
used the method of Wohlgemuth in determining amyloclastic activity. 
They stipulate a definite color for the end point of the iodine reaction, 
using the Milton Bradley color chart as given by Mullikin in his 
"Identification of Pure Organic Compounds." 

If the iodine method is to be used it appears probable that the most 
accurate results can be obtained by following the technic described 
by Sherman and his various co-workers. 

Copper Reduction Method for the Determination of Reducing Sugars 
Formed by the Action of Diastase. 

Although Kjeldahl showed that the reducing sugars formed by 
the action of diastatic enzymes upon an excess of starch was a 
measure of their activity, no great advance was made in the exact 
valuation of diastase preparations until 1886, when Lintner modified 
Kjeldahl's method. He first prepared a soluble starch possessing 
rather definite characteristics. He was then enabled to base his calcu- 
lations of diastatic power upon the production of a constant quantity 
of maltose formed by the action of the malt extract upon a definite 
amount of soluble starch. His method consisted of measuring 10 cc. 
of a 2 per cent soluble starch solution into a series of ten test tubes, 
to each tube he added a slightly different amount of malt extract. 
After digesting exactly one hour at 21 "^C, 5 cc. of Fehling solution 
is added to each tube and the whole series of tubes are placed in 
boiling water for ten minutes, and then examined to determine the 
first tube in which all the copper was reduced. Lintner prepared a 
diastase, of which .12 mg. was able to produce the required amount 
of maltose, under the above conditions, to completely reduce 5 cc. of 
Fehling's solution. To this preparation Lintner gave the value of 
100 and the power of the samples were calculated as inversely pro- 
portional to the amount of sample required to produce this fixed 
amount of reducing sugar. 

Ford (1904) prepared a soluble starch from various sources and 
after careful purification, similar to the Lintner method, concluded 
that no dift'erence was attributable to the source of the starch. He 
specified that sokible starch should be neutral to rosolic acid in order 
to give concordant results b}- the Lintner method. 

16 



Ford and Guthrie (^Iy08j expressed ani3lolylic activity as grains of 
maltose produced by a filtered extract of one gram of mashed barley 
in an excess of soluble starch for one hour at 40° C. 

Several other copper reduction methods have found their way into 
use since Lintner established his method for determining diastatic 
activity by measuring the reducing sugars formed. The most notable 
is that of Sherman. Kendall and Clark (1910). This method consists 
of placing different amounts of enzyme e. g., 0.2, 0.5, 0.8, and 1.0 
mgms. into four erlenmexer flasks, to this is added 100 cc. of a 2 per 
cent soluble starch and the whole digested 30 minutes at 40°C. At 
the end of this time, 50 cc. of Fehling solution are added and the flasks 
are immersed in a boiling water bath for 15 minutes. The reduced 
CU2O is then quickly filtered and determined gravimetrically. They 
supply a table which gives diastatic activity of the preparation called 
the "new scale.'' 

Influence of Temperature on Diastatic Activity. 

In all of the earlier work high temperatures were employed, that 
is between 40°-60°C. Marker found that at 60° four molecules of 
starch formed three of maltose and one of dextrin, while at 65° the yield 
was lowered and only one molecule ol maltose and one of dextrin were 
formed from two of starch. It is quite ob\ious from our present day 
knowledge that tlie optimum temperature for diastatic activity lies 
between 63°-65°C., which would explain why Marker got a decreased 
production of maltose at 65 °C. 

Brown and Heron (1879) found that previous heating of a malt 
extract decreased its activity to a great extent. They found that it 
showed very litle activity when heated above 66° C. 

Vernon (1901-1902) gives the optimum temperature for the activity 
of diastase as 35 °C. with continued activity up to 65 °C. 

Kendall and Sherman (1910) find the optimum temperature of puri- 
fied amylases to be 40° in the presence of salts and a trace of alkali. 
They find that between 20°-40° the activity is about doubled for each 
10° increase in temperature with a considerable falling off in rate of 
increase of activity betwen 40°-50°, where the maximum activity 
was found. 

Influence of Acids, Bases and Salts on Diastatic Activity. 

The effects of acids and bases upon diastatic activity has received 
a great deal of attention, some investigators claiming that an acid 
medium was the most favorable, while others advocate a neutral 
medium, for the optimum activity of the enzyme. 

Baswitz (1878-1879) and Mohr (1903) found that when carbon 
dioxide was passed through the reacting medium, a great increase in 

17 



diastatic activity took place. Detmer (1882) reached the same con- 
clusion and noted that small amounts of citric, phosphoric and hydro- 
chloric acids had an activating effect. Reychlcr (1889) found that 
KH.,P04 had a stinndating effect, while Eft'ront noted that HCl, 
HF, H.SO^ and pliosphoric acid as well as KH^PO^ had a very 
favorable action. Petit (1904) found an acid medium to be the best. 
Kjeldahl (1880) reported that H^SO^ when in a concentration of 
.005 N. increased diastatic activity, but was decidedly detrimental 
when .01 N. was used. The same concentrations hold for citric and 
acetic acids as reported by Schneidewind, Alcyer and Aliinter (1906). 

Hey] finds that KH^PO^ has an activating as well as a conserving 
action, while Hawkins (1913) reports that small additions of phos- 
phoric, acetic and tartaric acids increase the saccharogcnic power of 
the malt extract l)ut did not effect the amyloclastic activity. 

Chittenden and Cummins (1884), Duggan (1885). Lintner (1885), 
Ford (1904), Maquenne and Roux (1900) and Fernbach and Wolff 
(1907) are all agreed that a neutral medium is the most favorable for 
diastatic activity. 

Sherman and Thomas (1915) and Sherman, Thomas and Baldwin 
(1919) find that the optimum diastatic activity for various diastatic 
preparations, depending upon their source, is at a pH of 4.2-4.6. They 
report an optimum hydrogen ion concentration of al)Out pH 4.4 for a 
diastase prepared from malt extract, a pll of al)out 4.7 for the amylase 
of Aspergillus oryzae and a pH of 6.8-7.0 for pancreatic amylase. 
After reaching the optimum pll the activity falls off very sharply on 
the alkaline side. 

Many substances ha\e likewise been re])orted which activate dia- 
static })reparations such as asparagine, aspartic acid, diff'erent amino 
acids and proteins. Chief among the workers who have reported 
such results are Effront (1904). Ford (1904). Rockwood (1917). Sher- 
man and Walker (1921) and Sherman and Caldwell (1921). 

Very little work is reported on the action of alkalis upon diastatic 
activity. Brown and Heron report a decided falling off in diastatic 
activity when Ba(OH)o, KOH or NaOH is added to the medium. 
In fact, it has been the custom to use Alkalies to stop diastatic acti\it>- 
in solutions being analyzed. 

Effects of Proteolytic Enzymes. 

The action of proteolytic enzymes upou protein material is well 
understood today and needs very little mention. In regard to the 
action of proteolytic enzymes in flour, Ford and Guthrie were not 
able to show by any chemical means that tlicy existed in wdieat flour. 
However, when 1 per cent gelatin was added to the flour and then 

18 



allowed to solidify, proteolysis could be followed by the gradual 
liquification of the gelatin. 

Baker and Hulton could find no method to establish the presence 
of proteolytic enzymes in flour and therefore claim that any which 
may be present would have very little effect ujuju a dough. They did 
demonstrate, however, that yeast contains a very powerful protein 
splitting enzyme, as shown by the soluble nitrogen of two identical 
doughs, one with and the other without yeast. In the dough to 
which yeast had been added, they report 2.7 ])er cent soluble nitrogen 
as protein, while in the other dough they found onlv 1.9 per cent 
soluble nitrogen as protein. 

Hydrogen Ion Concentration. 

From the foregoing literature, it has been shown that acids, bases 
and salts are of the utmost importance in relation to the activity of 
diastatic ferments. Jessen Hanson has also shown that the optimum 
conditions for the baking of bread occur when the dough is at a 
hydrogen ion concentration of about pH 5.0. 

Bailey and Peterson (1921) find that when acid or alkali is added 
to buffered water extracts of flour, a characteristic curve is obtained 
which indicates very accurately the grade and baking qualities of a 
flour. Bailey and Collatz (1921) have shown that a remarkable 
parallelism exists between grade of flour and the electrical conduc- 
tivity of a water extract of flour when digested one hour at 25°C. 

Viscosity. 

Although Ford and Guthrie attempted to demonstrate the proteo- 
lytic action of enzymes in wheat flour, by digesting the flour in water 
at a set temperature by means of viscosity measurements, they report 
no success in this method. 

Ostwald and Liiers in a series of papers show that different mill 
grades of flours can be distinguished by means of a viscosimeter. 
From their data, the flours group themselves according to the degree 
of extraction in milling-. 

Quite recently Sharp and Gortner have demonstrated the efficiency 
of the viscosimeter in measuring differences in the imljibitional capac- 
ity of strong and weak flours when treated with \arious acids, bases 
and salts. They find that strong flours show a greater viscosity under 
the conditions of their experiments than do the weak flours. 

From the literature cited one may judge the amount of work 
expended upon the question of flour strength. Although the work 
of the last few years shows progress on this question, we do not have 
a single test which gives us an absolute criterion of flour strength 

19 



and it is still necessary tu fall back upon the baking tests to have 
a final answer to the question. 

II. EXPERIMENTAL. 
(a) The Problem. 

It has been shcnvn in the historical review that flour strength has 
been studied in a variety <>f ways. Two points of attack are outstand- 
ing, however, the work of W'ood and Hardy, Upson and Calvin, 
Gortner and Doherty, and Sharp and Gortner, who have concerned 
themselves with the physico-chemical properties of the gluten ; and 
of Baker and llulton. and I-'ord and Guthrie who have attacked the 
problem from the enzymic standpoint. In this Thesis we are con- 
cerned with enz}nie relationships. From tiie data presented by Ford 
and (lUthrie it would appear that the diastatic enzymes were of more 
importance than the proteolytic enzymes with regard to flour strength. 

Baker and Hulton indicate in their excellent work that the amy- 
loclastic enzymes A\ere perhaps the limiting factor in the production 
of maltose. I have taken up the i)roblem at this ])oint and am con- 
cerned with the efl:'ect upon the baking strength of wheaten flours 
when diastatic ferments are added to the dough. As the diastatic 
preparations a\ailable for the baker are in the form of malt flours 
and malt extracts which contain ])roteolytic enzymes, the problem 
is at once broadened to include the later as well as the starch splitting- 
ferments. 

(b) Material. 

The ])resent investigation was conducted with a commercial malt 
flour, a rei)re^entative malt extract, a commercial sample of wheat 
starch, and a series of seven wheat flt)urs of ditTerent grades milled 
from wheats grown in different regions of North America. All of 
the flour samples and malt preparations were sul)mitted to careful 
chemical analysis and in most cases were rechecked by other inves- 
tigators using the same materials. The buffer values of the flour 
extracts were also determined by the method of Bailey and Peterson 
in order to have additional data as to the grade of the flour. This 
data is given in Table VIT. 

Description of Materials Used in These Studies. 

The flours used in this inxestigation were flours milled from auth- 
entic samples of wheat, grown in different regions of North America 
under different climatic conditions. The A. O. A. C. methods were 
followed in analyzing the flours and malt preparations, with the 
results shown in Tables TV, V, and VI. A description of the wheat 
flours is as follows : 

20 



Flour 1001 was milled from a sample of hard Kansas wheat from 
the crop of 1921. The baking tests showed it to be of good strength 
and the low ash content. Hydrogen ion concentration, in terms of 
pH, show^ it to be a patent of low extraction. The protein content, 
which is a trifle low, reflects directly upon its l)aking strength. 

TABLE IV. 

Analysis of Wheat Flours on Air Dry Basis. 



Flour Samples 






Protein 






Laboratory 


Moisture 


Ash 


(Nx5.7) 


Milling 


pH of Water 


No. 


Percent 


Percent 


Percent 


Grade 


Extract 


1001 


12.15 


.40 


11.34 


Patent 


5.816 


1002 


12.14 


.61 


13.00 


Clear 


6.052 


1003 


13.06 


.46 


8.83 


Patent 


6.002 


1007 


11.06 


.64 


14.12 


Clear 


6.103 


1008 


11.70 


.46 


15.32 


Patent 


6.133 


1009 


11.61 


.42 


13.81 


Patent 


5.981 


1011 


11.44 


.56 


10.77 


Patent 


6.050 



TABLE V. 
Analysis of Malt Flour on Air Dry Basis. 





Total 




Reducinf;- 


Sugars 




Sugars 


as Dex- 


Diastatic Value 


s Dextrose 


trose 


Degrees 


Percent 


Percent 


Lintner 



Malt Flour 

Laboratory Moisture Ash Protein 
No. Percent Percent Percent 

24 8.8 1.26 11.25 4.75 10.62 177.05 

TABLE VL 
Analysis of Malt Extract. 

Reducing Sugars Total Sugars Dias- 

Calculated Calculated as Pro- tatic 

Ex- Ash DeX' Dex- tein Value 

tract Moisture Per- Maltose trose Maltose trose Per- Degrees Specific 

No. Percent cent Percent Percent Percent Percent cent Lintner Gravity 

D 25.63 1.35 73.74 42.52 73.80 42.62 6.06 37.1 1.384 

Flour 1002 was milled from a sample of hard Kansas wheat. The 
high ash content and the pH of the w^ater extract indicate a clear 
flour. The baking tests show it to have a fair degree of strength. 

Flour 1003 is a patent milled from soft, white winter, Washington 
wheat. The ash content and the pH of the water extract indicate 
a patent flour, while the protein content and the baking tests show 
it to be an exceptionally weak flour. 

Flour 1007 is a clear flour milled from a sample of selected hard 
spring wheat grown near Calgary, Canada. The ash content and the 
]iH, of the water extract, show it to be a clear flour. Although the 
]>rotein content is high the baking tests show it to be of poor baking 
strength. 

Flour 1008 is a patent milled from selected hard spring Canadian 

21 



wheat, and shows up exceptionally strong in the baking tests. This 
flour, it would seem, is too strong for any ordinary baking purposes 
and would have to be blended with a weaker flour. 

Flour 1009 is a composite, commercial patent flour, milled from 
hard spring wheat for a select trade. The ash content and pH values 
show it to be a low extraction patent, while the baking tests show 
it to be a very strong flour. Flmir 1009 does not give the volume of 
loaf that flour 1008 does, but it produces bread with a better grain 
and texture. This flour is the only one of the series in which the 
origin of the wheat is not known. 

Flour 1011 is a patent flour milled from Ohio winter wheat. The 
baking tests show it to be of rather poor baking strength. 

TABLE VII. 

Hydrogen Ion Concentration after addition of Acid and Alkali 
to Flour Extracts as an Index of Buffer Value. 

Malf 

Flour No. 1001 1002 1003 1007 1008 1009 1011 Flour 



pH pH pH pH pH pH pH pH 



cc. N/50 
HCl 
Added 

12..S 2.519 2.958 2.654 2.894 2.514 2.510 2.789 3.904 

10.0 2.654 3.006 2.874 3.359 2.876 2.705 2.977 4.225 

7.5 2.925 3.320 3.144 3.799 3.192 2.992 3.210 4.428 

5.0 3.388 3.685 3.761 4.471 3.545 3.496 3.630 4.727 

2.5 4.150 4.666 4.623 5.158 4.502 4.272 4.426 5.166 

0.0 5.816 6.052 6.002 6.103 6.133 5.981 6.050 5.491 

(cc.N/50 

NaOH) 

2.5 7.371 6.931 7.726 6.<»38 7.499 7.792 7.048 6.071 

5.0 9.045 8.021 9.653 7.852 8,906 8.892 8.883 6.390 

7.5 9.755 9.146 10.557 8.926 9.609 9.91S 9.540 6.652 

10.0 10.253 9.772 10.877 9.535 9.919 10.448 10.177 6.888 

12.5 10.617 10202 11.022 10.062 10.249 10.769 10.464 7.136 

(c) The Methods. 

The Munson and Walker method for the determination of reducing 
sugars was used throughout the investigation for the estimation of 
sugar resulting from diastatic activity, and all the results are calcu- 
lated as dextrose. In the cases where proteolytic activity is deter- 
mined, the amino nitrogen method of Van Slyke, and the viscosity 
method of Sharp and Gortner were used. 

The Method of Determining Diastatic Activity. 

It was evident from the very first that the method of Lintner, for 
the determination of diastatic activity was out of the question, as 
were also the other methods which have since been developed. It 
was also evident that the raw starch of the flours was the natural 

22 



substrate of the enzymes and consequently should be used to dupli- 
cate, as far as possible, the changes taking place in the fermentation 
process. Many difficulties were involved and clarification of the solu- 
tion was necessary to obtain uniform results. The method finally 
adopted was one which was developed and perfected in this laboratory. 
It consisted of adding 3 cc. of l57o Na.jWO^ to the digestion mixture, 
transferring to a 200 cc. volumetric flask and then adding 20 drops of 
concentrated HgSO^ and filling up to the mark with water. After 
careful and thorough shaking the contents were transferred to centri- 
fuge tubes and whirled 5 minutes. The resulting clear supernatant 
liquid contained all the soluble sugars and was practically free of 
soluble protein as demonstrated by repeated Kjeldahl determinations. 
For further data, see report on diastatic enzymes of wheat flr)ur and 
their relation to flour strength. (Rumsey, 1922). 

In determining the diastatic activity of a malt preparation and the 
amount of soluble sugars produced from the flours by its action, the 
following method was used : Ten grams of flour were weighed out 
and transferred to a 400 cc. erlenmeyer flask, the specified amount 
of flour or malt extract was then added. One hundred cc. of water, 
previously brought to temperature, was then added and the whole 
was thoroughly mixed and placed in a water bath, for 1 hour at 27° C. 
The flasks were agitated every five minutes and at the end of the 
digestion period the contents of the digestion flasks were transferred 
to a 200 cc. volumetric flask (any starch particles adhering to the 
sides of the flask can be removed with a rubber policeman and a 
stream of water from the wash bottle) and clarified as described 
above. After clarification 50 cc. aliquots were transferred to 400 cc. 
beakers and the reducing sugars determined by the Munson and 
Walker method. 

In determining the reducing sugars formed in the dough at various 
stages of fermentation essentially the same methods were followed. 
At the time specified, ten grams of dough are pinched off from the 
fermenting mass and rubbed up in a mortar with a little water until 
a homogeneous suspension is secured, this is then transferred to a 
250 cc. volumetric flask and the same procedure followed as outlined 
above. 

The Method of Determination of Proteolytic Activity. 

In the determination of proteolytic activity eighteen grams of flour 
(calculated on the dry basis) were weighed and transferred to a 500 
cc. erlenmeyer flask, malt flour or malt extract was added and 100 cc. 
of water, previously brought to temperature of digestion, was then 
added and the whole digested 4 hours at 40° C. The mixture was 

23 



then cuuled to 25' C, at the close of digestion, and poured into the 
cup of the MacMichael viscosimeter and the average of three readings 
taken. Then 5 cc. of N/1 lactic acid was added, the contents thor- 
oughly mixed, and the average of three successive readings taken. 
From the calibrated scale reading of the MacMichael Viscosimeter, 
which is denoted as M, any values such as centipoise or absolute 
viscosity can be determined by calculation. 

Method of Determining Buffer Value of Flours. 

In the determination of the buffer values of the flours, 80 grams 
of flour are weighed into a 2 liter flask and 400 cc of water added. 
The whole is well shaken up to get rid of any lumps and digested 
1 hour at 25 °C. The digestion mixture is then centrifuged to throw 
down the suspended matter. Aliquot portions of 25 cc. volume were 
created with 2.5. 5.0, 7.5, 10.0 and 12.5 cc. respectively of N/50 HCl 
or NaOH, then brought to a volume of 50 cc. and the hydrogen ion 
concentraticMT determined by the use of the Bailey electrode and a 
Leeds and Northrup ty])e K potentiometer in conjunction with an 
N/10 KCl calomel electrode and a flowing junction of saturated KCl. 

Method for Determination of Gas Producing Capacity of Flour. 

In determining tlie gas i)roducing capacity of a flour, twenty grams 
of flour are weighed out and transferred to a wide mouthed bottle. 
T(j this is added .5 grams of fresh yeast suspended in 20 cc. of dis- 
tilled water and the whole is thoroughly stirred and the bottle stop- 
pered with a cork containing a delivery tube. The bottles are then 
put in a water bath kept at 3>7°C. and the liberated gas is measured 
in an inverted cylinder under concentrated brine. Readings are taken 
every half hour. When malt extract is added it is first incorporated 
with the yeast and water and added to the flour in this way. 

Baking Tests. 

The .baking tests were carried out according to the standard formula 
adopted by the American Instiutc of Baking with one exception and 
that consisted of leaving out sucrose in one set of baking experiments. 
The formula of the dough was as follows: 

Flour ^•<23 grams 

Water 173 grams( varit-d ckpLiKling upon absorption of the flour) 

Yeast 10 grams 

Sugar 10 grams 

Salt 5 grams 

Lard .' 6.5 grams 

This formula was corrected for the sugar content of the malt flour 
and malt extract used, the total sugar content being always equivalent 
to that stated in the formula. The doughs were mixed with a small 

24 



bench mixer, fermented and baked under as accurately controlled 
conditions as possible. After the baked loaves were withdrawn from 
the oven they were placed in a cabinet to cool and were weighed 1 
hour after baking. The volumes of the bread were taken the next 
day and each loaf was then cut and judged for grain, texture, color, 
flavor and odor. In the case where the development of sugar forma- 
tion was followed during the course of fermentation, a double portion 
of dough was mixed and then divided. The samples for anlysis were 
taken from one portion of the dough while the other was baked, and 
then judged as were those described above. In these experiments 
dough was fermented, with and without yeast, to estimate the total 
production of sugar and that used by the yeast in normal fermenta- 
tion. The weight and temperature of the dough were taken at stated 
periods of fermentation. 

Certain changes in the hydrophyllic properties of the gluten col- 
loids of this series of doughs, as measured by the viscosity of dough 
suspensions in water, were followed by Mr. P. F. Sharp, and will 
be reported by him in a separate paper. 

(d) Influence of Varying Conditions on Diastatic Activity. 
Determination of the Optimum pH for the Amylase of Malt Flour. 

This analysis was made to determine what relation existed between 
the optimum pH of dough and the optimum pH for the maximum 
production of maltose by the malt flour used. Sherman and his col- 
laborators determined the optimum pH for a purified malt amylase 
and it was thought of interest to know how a commercial preparation 
behaved in this regard. The process of manipulation varied slightly 
from that described above for the determination of buffer values, so 
will be described at this time. Ten grams of malt flour containing 
both the enzyme and the raw-starch substrate were weighed out into 
a flask. Enough water was then added so that when the mixture was 
brought to the desired pH by acid or alkali, the total volume of liquid 
added would be 50 cc. The mixture was then digested for one hour 
at 25°C in an accurately regulated water bath. After digestion the 
whole was centrifuged and 25 cc. (half of sample) was pipetted into 
a 200 cc. volumetric flask, 2 cc. of 15% Na.,WO, were added and thor- 
ughly shaken, and 20 drops of concentrated H^SO^ is added and made 
up to the mark. The preparation was centrifuged again and 50 cc. 
taken for reducing sugars as described above. The other portion of 
the unclarified extract was used to determine the pH values. 

The experimental data showing the influence of diastatic activity by 
change in pH are given in Table VIII and illustrated graphically in 
Figure 1. The optimum activity occurred at pH=4.20. 



TABLE VIII. 

Relation between hydrogen ion concentration (as pH) and the 

diastatic activity of malt flour as expressed in grams of 

dextrose per 1.5 grams of malt flour. 









Wt. of 


Wt. of 




Xormalitv 


cc. 




Cu,0 


Dextrose 


Dextrose" 


of Hcr 


used 


pH 


Grams 


f^rams 


Per Cent 


N710 


28.0 


1.988 


.1075 


.0527 


3.52 


N/10 


26.0 


2.099 


.1115 


.0548 


3.65 


X/10 


24.0 


2.139 


.1147 


.0563 


3,75 


N/10 


22.0 


2.437 


.1111 


.0545 


3.64 


N/25 


50.0 


2.572 


.1070 


.0525 


3.50 


N 25 


45.0 


2.745 


.1123 


.0552 


3.68 


N/25 


40.0 


2.970 


.1239 


.0611 


4.07 


N/25 


35 


3.156 


.1438 


.0714 


4.76 


N/10 


13.6 


3.224 


.1526 


.0759 


5.03 


N/10 


12.0 


3.420 


.1734 


.0868 


5.78 


N/25 


30.0 


3.499 


1828 


.0917 


6.12 


N/10 


11.0 


3.613 


.1937 


.0976 


6.51 


N/25 


25.0 


3.704 


.1935 


.0974 


6.50 


N'25 


22 ^ 


3.870 


.1954 


.1000 


6.67 


N/25 


200 


4.159 


.2238 


.1137 


7.58 


N 25 


15.0 


4.260 


.2305 


.1173 


7.82 


N/25 


12.5 


4.542 


.2224 


.1129 


7.52 


N/25 


10.0 


4.621 


.2190 


.1111 


7.41 


N/25 


5.0 


5.069 


.2081 


.1051 


7.01 


NaOH 


0.0 


5.548 


.1827 


.0917 


6.11 


■^^" IN/25 


5.0 


6.069 


.1670 


.0834 


5.56 


N/25 


10.0 


6.4S9 


.1548 


.0769 


5.12 


N/25 


15.0 


6830 


.1438 


.0713 


4.75 


N 25 


20.0 


7.359 


.1336 


.0661 


4.41 


N/25 


25.0 


7.871 


.1230 


0605 


4.03 


N 25 


30.0 


8.419 


.1196 


.0589 


3.93 


N/25 


35.0 


8.920 


.1180 


.0581 


3.87 


N 25 


40.0 


9.306 


.1163 


.0572 


3.81 


N/25 


-1." 


9.649 


.1090 


.0535 


3.57 


N 2- 


: -^ , 


9.991 


.1028 


.0503 


3.37 



Influence of Time of Digestion on the Diastatic Activity of Malt Flour. 

The influence of time on the activity of malt flour was investigated 
to ascertain at what point, or length of time, the activity would 
decrease. Heyl has noted that at hrst the reaction is logarithmic, but 
deviates as the products of hydrolysis accumulate. This particular 
experiment was planned in order to find the optimum length of time 
for future periods of digestion. It developed that at the end of eight 
hours the reaction was proceeding at about the same rate of speed 
as that of one hour so it was decided to make one hour the standard 
period of all digestions. 

The efrects of time of digestion up to eight hours is given in Table 
IX and presented graphically in Figure 2. 



26 




Figure 1. 

Effect of pH on the activity of diastase in malt f^our expressed as 
grams dextrose per 1.5 grams malt flour. 

TABLE IX. 

Effects of time of digestion on diastatic activity as expressed in grams 
of dextrose per 10 grams of malt fiour. 







\\"t. of Dextrose 




Time of 




per 10 erams 




Digestion Weight of Cu:0 


of flour 


Dextrose 


Hour? Grams 


Grams 


Per Cent 


0.00 


1112 


.3844 


3.86 


.25 


1462 


.5128 


5.13 


.50 


1622 


.5752 


"> "^^ 


1.00 


1962 


.6992 


7.00 


1.50 


2230 


.7992 


8.00 


2.00 


2457 


.8848 


8.85 


3.00 


2931 


1.0696 


10.70 


4.00 


3331 


1 2296 


12.30 


5.00 


3650 


1.3608 


13.61 


6.00 


3991 


1.5024 


15.03 


7.00 


4264 


1.6200 


16.20 


8.00 


4560 


1.7480 


17.48 



27 



IMO 










































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1 
1 



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Figure 2. 

Diastatic activity as influenced by time of digestion, expressed in 
grams of dextrose per 10.000 grams of malt flour. 



Effect of Temperature Upon the Activity of Diastase. 

Practically all of the work in this investigation was carried out at 
27^C, which is the temperature of fermentation used in the bake shop; 
however, it was necessary to find the optimum temperature of the 
diastase in the malt flour, as Sherman notes that 40° is the optimum 
temperature with a maximum at 55° for pancreatic amylase. Most 
of the investigators quoted above found 65°C to be the optimum for 
malt amylase, while Swanson and Calvin found 62.5°C to be the 
optimum for wheat diastase. It was thought to be of interest to 
determine at what temperature the diastase in malt flour exerted its 

28 





























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/" 


"^ 


*>- 


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/ 










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1 
























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i.ooa. 








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MX 




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1 

1 







lempcroTi/re 

Figure 3. 

Effects of temperature upon the activity of diastase in malt flour when 
10.00 grams are digested one hour at various temperatures. 

TABLE X. 

Effect of temperature upon the activity of diastase in malt flour when 
10 grams are digested for 1 hour at various temperatures. 







Seri 


es 1 

Wt. of 
Dex- 






Serie 


Wt. of 
Dex- 




Temp. 






trose 








trose 




of 






per 10 








per 10 




Diges- 




Wt. of 


Grams 


Dex- 




Wt. of 


Grams 


Dex- 


tion 


Wt. of 


Dex- 


Malt 


trose 


Wt. of 


Dex- 


Malt 


trose 


Decrees 


Cu:0 


trose 


Flour 


Per 


CihO 


trose 


Flour 


Per 


C 


Grams 


Grams 


Grams 


Cent 


Grams 


Grams 


Grams 


Cent 


27 


.1279 


.0559 


.5590 


5.59 


.1280 


.0559 


.5590 


5.59 


40 


.1951 


.0867 


.8670 


8.67 


.1940 


.0862 


.8620 


8.62 


45 


.2305 


.1034 


1.0340 


10.34 


.2320 


.1041 


1.0410 


10.41 


50 


.2922 


.1333 


1.3330 


13.33 


.2914 


.1329 


1.3290 


13.29 


55 


.4144 


.1960 


1,9600 


19.60 


.4136 


.1956 


1.9560 


19.56 


60x 


.5704 


.2598 


2.5980 


25.98 


.5692 


.2592 


2.5920 


25.92 


65x 


.6030 


.2759 


2.7590 


27.59 


.6018 


.2754 


2.7540 


27.54 


70x 


.5760 


.2626 


2.6260 


26.26 


.5762 


.2627 


2.6270 


26.27 



X Aliquots of one-half the usual quantity were used, and the resulting values 
multiplied by two. 

29 



maximum effect. The procedure was as follows: 10 grams of maU 
flour were weighed out and digested at temperatures of 27°, 40°, 45*. 
50°, 55°, 60'. 65 , and 70 for one hour with 100 cc. of water (previ- 
ously brought to temperature). After clarifying and bringing to a 
volume of 250 cc. a 25 cc. aliquot was taken for reducing sugars and 
determined by the Munson and Walker method. These data are 
given in Table X and illustrated graphically in Figure 3. 

Effect of Concentration of Diastase on Hydrolysis of Starch 
in Wheat Flour, 

As the concentration of diastatic ferments added to the dough is of 
great importance, the effects of dift'erent concentrations of malt flour 
up to 50% were tried by mixing definite proportions of wheat and 
malt flour and digesting it at 27^ C. for one hour. It was necessar>' 
to first determine the amounts of dextrose formed when different 
amounts of malt flour were digested separately in order to apply cor- 
rections for the autolysis of malt flour in the succeeding experiments 
with the wheat flour. This data is given in Tables XI and XII, and 
presented graphically in Figure 4. 

TABLE XI. 
Autolysis of malt flour at 27 "C. for one hour. 

Amount of Mah Flour 

used, grams 0.25 

\Vt. Cu,0, grams .0337 

\Vt. Dextrose, grams .0142 
Dextrose, per cent 5.68 

TABLE XII. 

Relation of concentration of malt flour to the production of reducing 
Sugars from wheat flour when digested 1 hour at 27 "C. 



0.50 


0.75 


1.0 


1.25 


.0744 


.1080 


.1519 


.1980 


.0320 


.0469 


.0668 


.0852 


6.40 


6.25 


6.68 


690 









Dextrose 










weight of 


corrected 






Proportion 


\Vt ofCu.O 


Dextrose 


for dex- 






of wheat 
flour to 


per 2.5 


formed per 
2.5 gms. 


trose of 


Malt 


Dextrose 


malt flour 


gms. flour 


flour 


malt flour 


flour 


formed 




Grams 


Grams 


Grams 


Percent 


Percent 


10:0 


.0944 


.0408 


.0408 


0.0 


1.63 


9:1 


.2167 


.0968 


.0826 


10.0 


3.67 


8:2 


.2583 


.1168 


.0848 


20.0 


4.24 


7:3 


.2918 


.1331 


.0862 


33.3 


4.92 


6:4 


.3030 


.1387 


.0719 


40.0 


4.80 


5:5 


.3279 


.1512 


.0760 


50.0 


6.06 



30 



u 


1 J ; : i ■ 1 




; y^ 


S6 




: ^ 




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f^KCnf fft/T Flour 

Figure 4. 

Relation of increasing concentration of malt flour to the production of 
reducing sugars, when wheat flour is digested one hour at 27 "C. 

Effects of Increasing Amounts of Malt Flour When Digested With a 
Constant Quantity of Wheat Flour. 

Table XII shows the ettects of large quantities of malt flour upon 
wheat starch, but the amounts are out of all proportioii to that used in 
practice. In the experiments following, the sugar producing capacity 
of the malt flour was measured upon a series of seven flours and a com- 
mercial wheat starch with concentrations varying from 0.2 percent to 
5.0 percent. 

The experimental data showing percent dextrose produced, when 
10 grams of flour are digested with 0.02 - 0.50 grams of malt flour, is 
given in Table XIII and illustrated graphically in Figure 5. 



31 



TABLE XIII. 

Percent dextrose produced from 10 grams of flours 1001, 1002, 1003, 

1007, 1008, 1009, 1011, and a commercial wheat starch when 

digested with 0.02 - 0.50 grams of malt flour for 1 hour 

at 27°C. 









f: 


lour San 


iple Nuniber — 




— 


Com- 
mercial 


Ma 


lit Flour 
U.sed 


1001 


1002 


1003 


1007 


1008 


1009 


1011 


Wheat 
Starch 


Grams 


% 


% 


% 


% 


% 


7o 


% 


% 


% 


.0000 


0.00 


1.85 


1.34 


.40 


.94 


2.2S 


1.36 




.18 


.0200 


.20 










2.35 


1.72 






.0250 


25 


2.15 


1.50 


.74 


1.15 








.32 


.0400 


.40 












1.80 






.0500 


.50 


2.11 


1.64 


.88 


1.71 


2.38 




.85 


.42 


.0600 


.60 












1.95 






.0750 


.75 


2.47 


1.74 


1.03 


2.04 


2.48 






.50 


.0800 


.80 












1.98 






.1000 


.99 


2.59 


1.85 


1.09 


2.10 


2.54 


2.12 


1.00 


.59 


.1200 


1.19 












2.19 






.1250 


1.24 


2.69 


1,94 


1.14 


2.32 






1.06 


.62 


.1500 


1.48 


2.77 


2.04 


1.20 


2.50 


2.56 


2.28 


1.14 


.69 


.1750 


1.72 










2.58 


2.46 


1.16 




.2000 


1.96 


2.90 


2.11 


1.36 


2.80 


2.62 


2.62 


1.47 


.84 


.2250 


2.20 












2.64 






.2500 


2.44 


3.02 


2.21 


1.40 


2.95 


2.63 


2.67 


1.51 


.91 


.3000 


2.91 


3.15 


2.31 


1.49 


3.21 


2.72 


2.84 


1.56 


1.07 


.4000 


3.84 


3.34 


2.48 


1.64 


3.35 


2.83 


3.06 


1.72 




.5000 


4.76 


3.47 


2.61 


1.73 


3.7i 


3.03 


3.24 


1.93 


1.53 



The Production of Reducing Sugars in the Dough During 
Fermentation. 

The production of reducing sugar.s in an actively fermenting dough 
and its subsequent use by the yeast was followed at various stages of 
fermentation This necessitated running two parallel doughs, one 
having the required amount of yeast, while the other had no yeast 
added but identical in every other respect. After mixing the dough, a 
ten gram sample was pinched off, shaken to a homogeneous suspen- 
sion, clarified and the reducing sugars determined. At stated periods 
10 gram samples were taken and submitted to analysis, as after the 
mix. Twice the usual quantity of each dough (with and without 
yeast) was mixed, divided into two equal parts and the samples taken 
from one pijrtion only in order to have one dough to l^ake in the 
usual way. 

32 























































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Gromj Mall Flour 

Figure 5. 

Percent of dextrose formed from a series of flours when digested one 
hour at 21° Q. with various amounts of malt flour. 

Tables XIV-XXII give this data on three flours representing a 
strong type Canadian patent flour (1008), a decidedly weak Pacific 
Coast patent flour (1003), and a clear flour from Kansas (1002). Each 
flour was mixed with varying amounts of diastatic preparations as fol- 
lows. 1.5 percent malt flour, 4.0 percent malt flour and 3.0 percent 
malt extract. Controls with no diastase were included in each series. 
No sugars were added to any of the doughs except that amount con- 
tained in the malt flour and malt extract used, but this amount was 
deemed negligible in its eflfect upon frementation. 

33 



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Productions of reducing sugars in the panary fermentation of fllour 
1008 with different concentrations of malt flour and malt extract. 
(Full lines indicate doughs to which yeast was added, dotted lines no 
yeast dough). 



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Figure 7. 
Production of reducing sugars in the panary fermentation of flour 
1003 with different concentrations of malt flour and malt extract. 
(Full lines indicate doughs to which yeast was added, dotted lines 
no yeast dough). 

40 





























































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Time V Feimenhtm 



Figure 8. 

Productions of reducing sugars in the panary fermentation of flour 
1002, with different concentrations of malt flour and malt extract. 
(Full lines indicate doughs to which yeast was added, dotted Hnes 
no yeast dough). 

Determination of Proteolytic Activity as Measured by the Fall in Vis- 
cosity of Flour-water Suspensions, when Digested with 
Diastatic Preparations. 

The action of proteolytic ferments upon gluten is accompanied by 
a breaking down of the protein material into simpler compounds srch 
as protesoses, peptones and amino acids. As the latter can be easily 
measured by the amount of free amino nitrogen, it was thought th^t 
an increase in amino nitrogen measured by VanSlyke's method, would 
be a measure of the amount of proteolysis taking place. This, how- 
ever, was not the case as no appreciable difference in amino nitrogen 
could be detected when the flour was digested with and without malt 
preparations. 

41 



It is well known that dough becunies slack or less viscous during 
the fermentation period. This has been attributed to the action of the 
l)roteolytic enzymes of the }e3st and partially to those in n.ialt pr<"p- 
arations. ford and Guthrie were unable to demonstrate proteolytic 
enzymes in flour by means of the viscosimeter and concluded that any 
which were present would have little or no effect upon the gluten dur- 
ing fermentation. Sharp and Gortner have shown that the hydration 
capacity and the quality of the gluten can be accurately determined by 
the use of the viscosimeter. They have shown that the viscosity of a 
flour-water suspension is tremendously increased and increases to a 
well defined maximum by the addition of lactic acid in small amounts, 
while the further addition of lactic acid causes no appreciable change 
in viscosity value. The suspensions of various flours in water dif- 
fered slightly in initial viscosity and it was only after the addition of 
the lactic acid that these differences were increased to such an extent 
as to make the results of significance. Under the conditions of Sharp 
and Gortner's experiments the addition of 5 cc N/1 lactic acid was 

TABLE XXV. 

The measurements of proteolytic activity in a flour-water suspension 

showing the effects of different periods of digestion (1-5 hours) 

at 30°C. with 100 cc of water and concentrations of malt flour 

(2 and 4 percent) on viscosity (degrees MacMichael) 

with the addition of various amounts of 

lactic acid. 

Viscosity in degrees MacMichael on addition of N/1 lactic acid. 
N 1 Lactic Acid 



added 


cc 

Malt 
Flour 


. . .0.0 


0.5 


1.0 


1.5 


2.0 


3.0 


5.0 


Digestion used 


Degr's 


Degr's 


Degr's 


Degr's 


Degr's 


Degr's 


Degr's 


Hours 


Percent 


M 


M 


M 


M 


M 


M 


M 


1 


0.0 


33 


83 


119 


134 


144 


152 


160 


2 


0.0 


41 


107 


139 


151 


156 


161 


167 


3 


0.0 


36 


86 


120 


133 


139 


148 


156 


4 


0.0 


38 


86 


117 


129 


136 


144 


160 


5 


0.0 


34 


83 


115 


127 


133 


141 


153 


1 


2.0 


35 


86 


126 


135 


141 


147 


151 


2 


2.0 


29 


70 


101 


115 


122 


130 


135 


3 


2.0 


29 


67 


96 


108 


116 


122 


127 


4 


2.0 


30 


69 


97 


106 


112 


118 


122 


5 


2.0 


31 


70 


95 


104 


109 


115 


120 


1 


4.0 


32 


71 


104 


119 


127 


134 


138 


2 


4.0 


25 


57 


88 


101 


109 


117 


121 


3 


4.0 


22 


51 


77 


90 


97 


105 


111 


4 


4.0 


24 


48 


72 


83 


88 


96 


100 


5 


4.0 


22 


45 


68 


77 


83 


90 


96 



42 



sufficient to bring any flour to the point of highest viscosity ; accord- 
ingly their method was followed to see whether any differences could 
be detected when flour was digested alone and when digested with 
malt preparations and if any differences which existed in the flour- 
water extracts could l)c increased by the addition of lactic acid. 

A few preliminary experiments in which flour was digested with and 
without malt preparations for different periods of time showed that 
the flour-water suspensions of the different flours varied very little in 
their initial viscosities, thus showing why Ford and Guthrie were un- 
successful in measuring proteolysis by means of the viscosimeter. 
With the addition of lactic acid, however, great differences were notice- 
able between the different flours and the results justified the following 
experiments. 































































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Time ^ Dioestion m Hour> 

Figure 9. 

Effect of the proteolytic enzymes, contained in malt flour, upon 
wheat flour as measured by the fall in viscosity when digested 1 to 5 
hours at 30°C. With lactic acid added. 



43 



The e::TcCi ■- i time ^'f d!g"e?Lion was nrsi tried to determine the op- 
tiniuni tin-e .f digestion. The experiment consisted of digesting flour 
lOC'l ;a strong Kansas patent) for 1 to 5 hours with 100 cc of water, 
and reoeating with the same liour after adding 2 and 4 percent of malt 
!l:ur. After digesting fcr the stated period oi tinne the flour-water 
suspensions were transferred to the cup of the MacMichael viscosi- 
meter and the average oi three readings taken. X 1 lactic acid was then 
added in sm.all amounts up to 5 cc. The mixture was thoroughly 
stirred aJter each addition of acid and the average of three viscosity 
readings taken. 

The ex-- erin: ental data of this series when digested at 30' C. is given 
in Table XX\' and the data in the last column < -i this Table is shown 
graphically in Figure 9. 

It a^vcars fr:n". this table that four h'-ur digestion is ample tinie to 
secure evidence of prcaeolytic action, and this time of digestion was 
used in all subsequent wc-rk. Also the temperature of digestion was 
increased from 30" to 40' in 'irder to procure greater activity of the 
enzyn-.es. 

The data given in Table XX\T and illustrated graphically in Figure 
10 was obtained by digesting a series of flours . 1001-1002-1003-1007 
and ICOS.' with varying amounts of malt flj^ur 1.0-1.5-2.0-2.5-3.0 and 
4 Dcrcent > with iCO cc of water at -rO'C. and determining the viscosity 
after adding 5 cc of X 1 lactic acid. Table XX\'II and Figure lOA 
shrw similar data v/hen malt extract was used. 

It has ':een sh I'V.m. in the literature cited that even minute traces of 
salts have a marked affect upon the imbibition capacities Cif gluten. 
It v.-as. therefore, thought advisable l'j see how the viscosity of the 
f.:ur reacted v.-hen treated as above but with the salts of the flour and 
n:alt n:ur washed -ut after the digestion period. The precedure wa^ 
as foll;v.-s: The equivalent of IS gram.s of water-free flour 1002, was 
weighed int.^ each of ^hc nasks and digested with 0.0. 1.0. 1.5. 2.0, 2.5. 
3.0 and 4.0 percent m.alt flour in lC>Ci cc of v/ater at -K)'C. After for.r 
hours 910 cc of distilled water v.-ere added, the vrhole well shaken and 
centrifuge:. The suternatant liquid v."as decanted and the flour 
residue shaken up with v.-ater and after complete disintegration made 
uo to ICO cc. olaced in visC'Osim.eter cup. and the readings taken after 
adomg varym.g am. ^'Unts oi lactic acio. 

The data sh :'V,-ing the viscosity in the presence of lactic acid of flour 
lC0i2 v.-ith the salts v.-ashed out is given in Table XX\'III. 

In order to dem.onstrate still further that the salt content of the 
added m,alt flour did not vitiate the results obtained in Table XX \ I 
v.-hen f.i'ur f:r exam.ple i 1C02 v/as digested 4 hours with 4 percent 

44 



TABLE XXVI. 

Effect of varying amounts of malt flour upon the viscosity of 18 grams 

(calculated on the dry basis) of wheat flour when digested for 

four hours at 40°C. with 100 cc of water (5 cc N 1 lactic 

added in each instance). Viscosity readings in 

degrees MacMichael. 



% Malt 
















Flour Used. . 


.. 0.0 


1.0 


1.5 


2.0 


2.5 


3.0 


4.0 




Degr's 


Degr's 


Deer's 


Degr"s 


Desr's 


Degr's 


Desr 


Flour Xo. 


^f 


M 


M 


M 


M 


-M 


M 


1001 


145 


123 


119 


114 


112 


105 


93 


1002 


150 


126 


118 


109 


97 


99 


81 


1003 


50 


44 


42.5 


40 


36.5 


37 


28 


1007 


124 


106 


103 


95 


97 


103 


88 


1008 


151 


136 


129.5 


117.5 


112 


109 


102 




nrztr' Ma.'T firfim.-m^'xr ior 



Figure 10. 

Changes in viscosity of fiour water suspensions when digested 4 
hours at 40 "C. with increasing amounts of malt flour and malt extract. 
as measured in degrees MacMichael with lactic acid added. (Full 
line curves flours digested with malt flour, dotted line curves those 
digested with malt extract). 



TABLE XXVII. 

Effect of varying amounts of malt extract upon the viscosity of 18 

grams (calculated to the dry basis) of wheat flour when digested 

4 hours at 40 C. with 100 cc of water. (5 cc of N/l lactic 

acid added in each instance). Viscosity readings 

in degrees MacMichael. 



% Malt 
















Ext. Used . . 


. ...0.0 


1.0 


1.5 


2.0 


2.S 


3.0 


4.0 




Degr'.s 


Dear's 


Decjr's 


Degr's 


Degr's 


Degr"s 


Degr's 


F"lour No. 


M 


M 


M 


M 


M 


U 


M 


1002 


145 


136 


131 


127.5 


120.5 


111 


106 


1003 


3/ 


51 


50 


48 


43 


38 


34 


1008 


ISO 


138 


134 


132 


127 


122 


112 



/5 
















! 




1 
1 


R 




1 i 


1 






6S 






1 
1 






« 












; ' . I 


J 

3e « 










<" 

.. «. 








i : , . 


1, 


"^ 








1 ' ^ 




s 




i 


1 ■ . ■ ' ' 


3 


1 






' ' i i 1 i 




*i 






K. r^- 








1 


1 

i 




« 








j\- 


"i 
















X 










LV--~ 






1 


1 


ji 
















1 


11 












1 ^ 


1 , 



Rrcenf Mair Pnjxtrahon Uied 



Figure 10-A. 

Changes in viscosity of water suspensions of flour 1003 when 
digested 4 hours at 40° with increasing amounts of malt flour and 
malt extract, as measured in degrees MacMichael. With lactic acid 
added. (Full line curve digestions with malt flour, dotted line curve 
digestion with malt extract). 



malt flour three samples of 1002 were weighed out, two were used as 
checks and to the third 4 percent malt flour was added. At the end 
of three and one-half hours 4 percent malt flour was added to ore of 
the checks and the mixture well shaken. At the end of 4 hours diges- 
tion, the viscosities of all three preparations were determined as usual. 
The data showing the viscosities of this experiment are given in 
Table XXIX, and leave no doubt but that proteolysis has taken place. 

TABLE XXVIII. 

The effect on the viscosity of flour 1002 with the salts washed out af- 
ter digesting with varying amounts of malt flour for 4 hours at 
40°C. with 100 cc of water (lactic acid added in each 
instance.) 

Lactic Acid C€. 0.5 cc 1.0 cc. 1.5 cc. 

Viscosity Viscosity Viscosity 

Percent Degrees Degrees Degrees 

Malt Flour M MM 

388 411 405 

1 299 328 340 
Z 295 312 315 

3 273 295 305 

4 245 263 271 

TABLE XXIX. 

Effect of three and one-half hour digestion without, and half an hour 

digestion with malt flour as compared to a 4 hour digestion with 

and without 4 percent malt flour upon the viscosity of 

suspensions in water. 

Culjic Centimeters N/1 Lactic Acid Added 0.0 5.0 

171 XT Degrees Degrees 

Flour No. |j 1^ 

1002 digested vvitiiout mah flour 38 145 

1002 digested with 4% malt flour 19 81 

1002 digested 3.5 hrs. without and 3 minutes witli 

4% malt flour 30 127 

Gas Production Capacity of Wheat Flour in Relation to Strength. 

Although Wood pointed out that the gas production capacity of a 
flour was an index to one of the factors in flour strength and Baker 
and Hulton pointed out that weak flours were low in liquifying' en- 
zymes, they did not sul)mit sufficient data to show that this was ac- 
tually the case. In the following work the gas producing capacities 
of a series of flours was determined, according to the method of Wood, 
and with and without the addition of malt extract. The flours selected 
consisted of two typically strong patent flours 1008 and 1009, two clear 
flours of fair baking strength 1002 and 1007, and one typically weak 

47 



patent flour 1003, milled from Washington wheat. The method fol- 
lowed was the same as that described in the methods under gas pro- 
duction. 

The data giving the cul)ic centimeters of gas produced from flours 
1002, 1003, 1007, 1008 and 1009 when fermented with and without the 
addition of 1 percent malt extract, is given in Table XXX, and illus- 
trated graj)hicall\' in Figures 11 and 12. 

TABLE XXX. 

Effect of added malt extract upon the gas producing capacity of flours 

1002, 1003, 1007, 1008 and 1009 when fermented with 2.5 percent 

yeast for four hours at 35°. 

No Added Malt Extract One Percent Malt Extract 

Flour No 1002 1003 1007 1008 1009 | 1002 1003 1007 1008 1009 

Time of ■ I 

Fermentation 
Hours 

0.5 
1.0 

1.3 
2.0 
2 =i 
3.0 
3.S 
4.0 
4.5 .128 I .. 208 

TABLE XXXL 

Changes in pH during the fermentation of the dough with values for 

flour extract and the extract of bread crumb. 

Baking Data. 



jas 


Gas 


Gas 


Gas 


Gas 


Gas 


Gas 


Gas 


Gas 


Gas 


cc. 


cc. 


cc. 


cc. 


cc. 1 


cc. 


cc. 


cc. 


cc. 


cc 


13 


14 


10 


10 


7 1 


14 


15 


12 


14 


8 


34 


M 


31 


27 


2H 


35 


34 


33 


36 


31 


62 


57 


58 


53 


60 1 


64 


59 


60 


66 


61 


98 


80 


92 




90 


99 


88 


94 


105 


101 


140 


95 


132 


148 


116 


141 


115 


134 


153 


125 


177 


104 


174 


202 


148 


180 


140 


178 


203 


153 


214 


117 


212 


250 


176 


I 218 


184 


22i 


252 


181 


245 


123 


243 




192 1 


254 


196 


270 




196 



Flour 


Ash 


Flour 


Mix 


1st Pch. 


pH Valu 
2nd Pch. 


es 

3rd Pch. 


After Pf. 


Bread 


No. 


7c 
















1001 


.40 


5.81 


5.33 


5.18 


5.09 


5.02 


4.79 


4.96 


1002 


.61 


6.05 


5.65 


5.25 


5.25 


5.24 


5.05 


5.2M 


1003 


.46 


6.00 


5.24 


5.16 


5.19 


5.37 


5.05 


5.16 


1004 


.83 


6.17 


5.91 


5.92 


5.87 


5.85 


5.80 


5.52 


1005 


.43 


5.84 


5.75 


5.40 


5.22 


5.23 


5.17 


5.29 


1006 


.38 


5.78 


5.70 


5.28 


5.22 


5.17 


5.03 


5 30 


1007 


.64 


6.10 


5.76 


5.63 


5.58 


5.59 


5.53 


5.. 58 


1008 


.42 


5.98 


5.33 


5.23 


5,17 


5.19 




5.25 



1011 .56 6.15 5.47 5.33 5.30 5.20 4.92 5.28 

Change in Hydrogen Ion Concentration of Fermenting Dough. 
As considerable data has been accumulated upon this series of flours 
it w\'is thought that the changes in hydrogen ion concentration dur- 
ing the fermentation period would be of considerable value, inasmuch 
as the speed of diastatic and proteolytic activity is influenced to such a 
great degree by changes in hydrogen ion concentration and many ir- 

48 















































































































\ 






















/ 






























f 














s^ 














/ 


/ 


K 


























1/ 


/ 














o 












//> 


/ / 






,mJ- 








^ 












//// 


^ ... 


























/ 


// . 


















Jt 








^ 


f 


























^ 


f 



























^ 


^ 

























Time ,•, Hci 



Figure 11. 

Gas producing capacity of flours 1002, 1003, 1007, 1008 and 1009, 

fermented with 2.57c yeast. 

































3a. 


























































2S0 












































/ 


/ 












100 
















// 




llfi- 










S' 














/ 


/, 


// 












1 /JO 














// 


























//> 


///. 
















/OOf 












w ■ ' 
















































■SO 








y 




























A 


/ 



























^ 


^ 

























Time m He. 






Figure 12. 
Effect of 1.9' added malt extract upon the gas producing capacity of 
flours 1002, 1003, 1007, 1008 and 1009, when fermented with 25.7^ yeast. 

49 



regularities in the data might be accounted for in this manner. The 
activity of yeast is also very much influenced by changes in the pH of 
its medium. The doughs were made from the same flours and in the 
same manner as that reported in the section on reducing sugar? 
formed during fermentation. The procedure consisted of taking 10 
grams of dough and shaking it up with 50 cc of water until a homo- 
geneous suspension was secured. The whole was then centrifuged 
and the pH of the supernatant liquid was then determined in the man- 
ner described above. Samples were taken at the mix, first punch, 
second punch, third punch and after proof. The pH value of the flour 
extract is also given as is that of a water extract of the finished bread. 
The data is shown in Table XXXT. 

In the past all chemical and physical data accumulated on th.e 
strength of flour has been accompanied by baking tests which in the 
final analysis have been the criterion of flour strength. Inasmuch as it 
was imperative to have accurate knowledge of the flours and the ef- 
fects of diastatic ferments, a series of baking tests was conducted in ad- 
dition to the baking tests made in studying formation of reducing 
sugars and the change in hydrogen ion concentration during the fer- 
mentation process. All baking tests were conducted as nearly alike as 
possible to secure comparable data. 

In the baking experiments conducted to test the elYects of added 
diastatic ferments upon wheat flours, the two diastatic preparations 
used throughcHit the entire work were employed, namely a malt flour 
and a malt extract. These were added in amounts of .5, 1.0, 1.5, 2.0, 
2.5, 3.0 and 4 per cent to each of a series of flours and the results are 
recorded as total lime of fermentation, valume of the loaf, color, grain 
and texture of the crumb, flavor and odor. The doughs were made in 
the manner described under the methods of baking tests and where 
fermentation is spoken of, total fermentation is meant including both 
the actively fermenting and proofing periods. 

The data shcnving the efl'ects of varying amounts of malt flour and 
malt extracts upon flours 1001. 1002, 1003. 1007, and 1008 is given in 
tables XXXII to XLI. 

TABLE XXXII. 
The effects of varying amounts of malt flour upon the baking qualities 

of flour 1001. 

Malt Flour % .Standard 5 1.0 \.S 2.0 2.5 3.0 5.0 

Fermentation Hrs. 5:00 4:56 4:52 4:48 4:44 4:40 4:36 4:32 

Sucrose added gms. 10.00 9.7Q0 0.580 9.370 9.155 8.940 8.730 8.310 

\Vt. of dough gm^. 513 515 516 517 517 521 .522 525 

\Vt. of loaf gms. 454 452 443 446 450 446 459 457 

\ol of loafer. 1870 1890 2020 1955 2030 2060 1910 2010 

50 



General Remarks : Loaves grade down with respect to grain and 
texture with each added increment of malt flour. Color of the crumb 
is markedly influenced by each increase of malt flour. The crumb was 
full of large gas holes which was probably due to localized yeast ac- 
tivity. Mean average temperature of fermentation was 81 °F., and the 
temperature of baking 440° F. Time of baking 26 minutes . 

TABLE XXXIII. 

The effects of varying amounts of malt extract upon the baking qual- 
ities of flour 1001. 

Malt Extract % Standard .5 1.0 1.5 2.0 2.5 3.0 5.0 

Fermentation Hrs. 5:00 4:52 4:48 4:44 4:38 4:34 4:30 4:26 

Sucrose added gms. 10.00 8.94 7.88 6.820 5.76 4.70 3.64 1.52 

Wt. of dough gms. 513 514 514 515 516 514 515 517 

Wt. of loaf gms. 454 457 450 448 447 450 453 457 

Vol. of loaf cc. 1870 1810 1880 1895 1925 1920 1970 1880 



General Remarks : The texture and grain was excellent through- 
out, but the loaf made with 1 percent malt extract seemed to have 
better grain than any other. Color was very good up to 3 percent of 
malt extract where increase in the malt extract darkens the color, or 
shade. The volume of the loaf also increases up to 3 percent malt 
extract and then drops. A very decided sweet honeyed flavor was im- 
parted to the bake which grew more pronounced as the percentage of 
malt extract increased. Mean temperature of fermentation 81°-82°. 
Time of baking 25 minutes. Temperature of baking 440°-430°C. 

TABLE XXXIV. 

The effects of varying amounts of malt flour upon the baking qaulities 

of flour 1002. 

Malt Flour % Standard .5 1.0 1.5 2.0 2.5 3.0 5.0 

Fermentation Hrs. 5:18 5:13 5:08 5:03 4:58 4:53 4:48 4:44 

Sucrose added gms. 10.00 9.79 9.58 9.37 9.155 8.94 9.73 8.31 

Wt. of dough gms. 519 523 525 528 529 530 532 535 

Wt. of loaf gms. 454 451 454 451 460 458 470 471 

Volof loaf cc. 1740 1890 2040 2010 2050 2010 2050 2030 

General Remarks : Color of crumb grades down very quickly with 
each addition of malt flour. Grain very much alike throughout while 
texture was even. Color of the crust improves wdth increase in malt 
flour; very good smell and good taste while malt flavor is not in evi- 
dence. Mean Temperature of fermentation 82°-83° and baking 470°F. 
Time of baking 24 minutes. 

51 



Standarc 


1 ,5 


1.0 


1.5 


2.0 


2.5 


3.0 


5.0 


5:19 


5:12 


5 :07 


5:02 


4:57 


4:52 


4:47 


4:42 


10.00 


8.94 


7.88 


6.82 


5.76 


4.70 


3.64 


1.52 


519 


514 


513 


513 


513 


514 


515 


521 


454 


441 


448 


445 


437 


436 


440 


447 


1740 


1900 


2050 


2010 


2010 


1980 


1060 


1940 



TABLE XXXV. 
The effect of varying amounts of malt extract upon the baking qual- 
ities of flour 1002. 

Malt Extract % 

Fermentation Hrs. 
Sucrose added gnis. 
Wt. of dough gms. 
Wt. of loaf ;;ms. 
Vol. of loaf cc. 

General Remarks : Color of crumb grades off a trifle as percentage 
of malt extract increases. Texture increases in fineness with increase 
in malt extract. Grain is decidedly improved with an addition of malt 
extract up to 2 percent and then falls off. Odor and flavor of malt ex- 
tract increases as the percentage of malt extract increases. No no- 
ticeable difference in the color of the crust between the various bakes. 
Mean temperature of fermentation S2'^-83° and Ixiking 479° F. Time 
of baking 25 minutes. 

TABLE XXXVL 
The effect of varying amounts of malt flour upon the baking qualities 

of flour 1003. 

Mah Flour % 

Fermentation Hrs. 
Sucrose added gms. 
Wt. of dough gms. 
VVt. of loaf gms. 
Vol. of loaf cc. 

(Jeneral Remarks: Standard loaf had by far the best color, which 
grades down very quickly with increase in malt flour. Loaf made 
with 0.5 percent malt flour possessed the best texture, flavor and odor. 
That with 1 percent malt flour had the best grain and those loaves with 
increased quantities grade off to a very coarse grain. Loaves were 
^oggy and heavy. Mean temperature of fermentation was 81° and 
that of baking 470° F. Time of baking 21 minutes. 

TABLE XXXVIL 

The effects of varying amounts of malt extract upon the baking quali- 

ities of flour 1003. 

Malt Extract % 

Fermentation Hrs. 
Sucrose added gms. 
Wt. of dough gms. 
Wt. of loaf gms. 
Vol. of loaf cc. 



.Standar( 


1 .5 


1.0 


1.5 


2.0 


2.'? 


3.0 


5.0 


4:42 


4:38 


4:34 


4:28 


4:24 


4:20 


4:16 


4:12 


10.00 


9.79 


9.58 


9.37 


9.155 


8.94 


8.73 


8.31 


511 


512 


515 


515 


513 


514 


521 


523 


468 


467 


468 


464 


457 


463 


469 


476 


1660 


1760 


1675 


1860 


1710 


1690 


1650 


1550 



Standat 


■d .5 


1.0 


1.5 


2.0 


l.S 


3.0 


5.0 


4:42 


4:44 


4:40 


4:36 


4:32 


4:28 


4:25 


4:21 


10.00 


8.94 


7.88 


6.82 


5.76 


4.70 


3.64 


1.52 


511 


504 


507 


504 


506 


508 


' 508 


492 


468 


460 


461 


454 


448 


453 


460 


448 


1660 


1 730 


1775 


1810 


1770 


1700 


1750 


1680 



General Remarks : Color was decidedly the best in the loaf made 
with 0.5 percent malt extract while the texture and flavor were best in 
that with 2.5 percent. Best grain was secured when 1.5 percent malt 
extract was used and seemed to run off as percent malt extract in- 
creased but nearly as bad as that of the malt flour. The malt flavor 
was not as pronounced as in the previous bakes when using malt ex- 
tract. Color of crust good throughout. Mean temperature of fer- 
mentation 81° and that of baking 525° F. Time of baking 20 minute.s 

TABLE XXXVIII. 

The effect of varying amounts malt flour upon the baking 
qualities of flour 1007. 



Malt Flour % 


Standai 


•d .5 


1.0 


1.5 


2.0 


2.5 


3.0 


5.U 


Fermentation Hrs. 


5:22 


5:15 


5:10 


5:06 


5:02 


4:58 


4:52 


4:47 


Sucrose added gms. 


10.00 


9.79 


9.50 


9.37 


9.155 


8.94 


8.73 


8.21 


Wt. of dough gms. 


















Wt. of loaf gms. 


















Vol. of loaf cc. 


1640 


1750 


1750 


1780 


1680 


1665 


1610 


1690 



General Remarks: 1.5 percent malt flour gave the largest volume, 
hnest texture, color and grain. This appeared to be the high point in 
quality, since all factors decreased as percentage of malt flour in- 
creased. Mean temperature of fermentation was 84° while that of 
baking was 470° F. Time of baking 22 minutes. 

TABLE XXXIX. 

The effect of varying amounts of malt extract upon the baking 
qualities of flour 1007. 



Malt Extract % 


.Standar 


■d .5 


1.0 


1.5 


2.0 


2.5 


3.0 


5.0 


Fermentation Hrs. 


5:22 


5:17 


5:12 


5:07 


5 :02 


4:57 


4:52 


4:47 


Sucrose added gms. 


10.00 


8.94 


7.88 


6.82 


5.76 


4.70 


3.64 


1.52 


Wt. of dough gms. 


















Wt. of loaf gms. 


















Vol. of loaf cc. 


1640 


1690 


1900 


1620 


1660 


18.^0 


1890 


1760 



General Remarks : The tise of 3 percent malt extract gives the best 
loaf for color, texture and grain and the best general appearing loaf. 
Malt extract increased the bloom, color of crumb and volume, through- 
out the bake. Mean temperature of fermentation was 84° while that of 
baking- was 470° F. The time of baking was 25 minutes. 

TABLE XL. 
The effect of varying amounts of malt flour upon the baking qualities 

of flour 1008. 
Malt Flour % 
Fermentation Hrs. 
Sucrose added gms. 
Wt. of dough gms. 
Wt. of loaf gms. 
Vol. of loaf cc. 

53 



Standar 


d .5 


1.0 


1.5 


2.0 


2.5 


3.0 


5.0 


4:53 


4:57 


4:52 


4:47 


4:42 


4:37 


4:32 


4:27 


10.00 


8.94 


7.88 


6.82 


5.76 


4.70 


3.64 


1.52 


524 


524 


525 


526 


630 


530 


531 


532 


450 


460 


455 


460 


467 


466 


467 


464 


2160 


2070 


2000 


2120 


2100 


1950 


1860 


1885 



General Remarks : The color of the crumb was affected by the addi- 
tion of malt flour as those preceeding. Best texture and grain was 
secured by the use of 1.5 percent malt flour. There was a very 
marked difference between tln»so loaves made with 1.5 and 2.0 percent 
in texture and grain. A distinct whcaty ^mell was n(,)ticed in the 
loaves having malt flour. The color and bloom were about alike. 
Temperature of fermentation was 83° and the temperature of baking 
480° F. Time of baking was 24 minutes. 

TABLE XLI. 
The effect of varying amounts of malt extract upon the baking quali- 
ties of flour 1008. 



Malt Extract % 


Standai 


■d .5 


1.0 


1.5 


2.0 


2.5 


3.0 


5.0 


Fermentation Hrs. 


4:53 


4:57 


4:52 


4:47 


4:42 


4:37 


4:32 


4:27 


Sucrose added gms. 


10.00 


8.94 


7.88 


6.82 


5.76 


4.70 


3.64 


1.52 


Wt. of dough gms. 


524 


513 


513 


514 


518 


521 


519 


525 


Wt. of loaf gms. 


450 


457 


457 


454 


450 


450 


453 


460 


Vol. of loaf cc. 


2160 


2020 


2040 


2070 


1995 


2130 


2180 


2130 



General Remarks : Color was good throughout the bake, the bread 
made with 3 percent had the best grain and texture. Added malt ex- 
tract gave the bread a slight sweet odor and taste. Bloom was even 
throughout the whole bake. Temperature of fermentation 83° and 
was baked out in 22 minutes at a temperature of 500° F. 

III. DISCUSSION. 
Changes in pH, Temperature, Time and Concentration and Their Ef- 
fects Upon the Activity of the Diastases Contained in a Com- 
mercial Malt Flour. 

As already noted in the historical review of the diastase literature, 
Sherman and his co-workers found that the pH for the optimum ac- 
tivity of diastase of different origins were not the same and it could 
hardly be expected that the diastases derived from different sources of 
barley would have the same activity at the same hydrogen ion concen- 
tration. A study of Table Villi, and Figure 1 show that the greatest 
activity of the diastases,- in the malt flour used in this investigation, 
was at a pH of 4.26, while that found by Sherman for a highly purified 
malt amylase was very close to a pH of 4.4 which shows relatively 
close agreement. It will be noted in Table XXXI, where the changes 
in hydrogen ion concentration of fermenting dough was followed, that 
in the later stages of fermentation the dough was, with two exceptions, 
at a pH of about 5.0. Although this is not at the optimum for dias- 
tatic activity, it will be noted from Figure 1 that the rate of reaction 
was very high at this point. This is of significance when we consider 
that the sugars formed in the later stages of fermentation are impor- 
tant factors in determining the size of the resulting loaf. 

54 



Table IX and Figure 2 show that the diastatic activity of the malt 
flour was practically constant over a period of eight hours digestion. 
A slight decrease in activity was noticeable as time of digestion pro- 
ceeds, but for all practical purposes, the rate of reaction showed a 
straight line when the quantity of dextrose formed was plotted against 
time. 

Table X and Figure 3 show that increase in temperature up to 
65°C. increased the activity of the diastase. From 27° - 45°C. the rate 
of reaction increased quite regularly with each increment of rise in 
temperature. Between 45° and 50° the rate was greatly increased, 
while between 50° and 60° the increase was very rapid, following 
quite closely the Vant Hofif and LaBelle law. After 60° the increase 
in activity was not so marked and the diastatic activity was apparently 
at a maximum at 65 °C., as a decline in activity was noted with further 
increase in temperature. Table XII shows that when the percentage 
of malt flour was increased from 0-50 per cent, the percentage of dex- 
trose formed from malt flour increased from 1.63 to 6.06 percent. This 
was calculated to show the quantity of raw starch converted to dex- 
trose. The greatest efifect of added diastase was in the first 10 percent 
of added malt flour, which gave an increase in dextrose from 1.03 to 
3.67 percent. 

The Effect of Diastatic Enzymes Upon Starch of Different Flours. 
The addition of diastatic ferments to wheat flours increased the re- 
ducing sugars, when digested at 27°C. for 1 hour. As the amount of 
diastatic ferment was increased a corresponding increase in reducing 
sugars was noted. All of the flours used in the experiment did not 
react in the same way to the addition of malt flour, as great dififerences 
were shown not only in the initial amounts of reducing sugars (1 
hour digestion without diastase) contained, but with an increase in 
malt flour more starch was converted in some flours than in others. 
In general, but for one exception, the weaker flours produce less re- 
ducing sugars than do the stronger, when digested with the same 
amounts of malt flour. From the data presented in Table XIII and 
Figure 5, it will be noted that the commercial wheat starch shows the 
least amount of initial reducing sugars and responds less to the 
action of malt flour than any of the wheat flours. Flour 1003, a 
decidedly weak Pacific Coast flour, is next in the series; it shows a 
slightly greater amount of initial reducing sugars (digestion 1 hour 
without added diastases) and responds a trifle more readily to conver- 
sion by the malt flour, than does the wheat starch. The next flour, 
1011, is a patent milled from a soft winter wheat and runs just a trifle 
higher in initial sugar content and appears to be more easily converted 
than flour 1003. These two flours, and the wheat starch constitute a 

55 



special group a> far as sugar content is concerned. W hile the wheat 
starch has no baking value, the other two, namely 1003 and 1011. 
showed very poor baking qualities. 

The patent flours of good baking strength are highest in the list of 
initial sugar content and with one exception produce under the action 
of diastatic enzymes, more dextrose than do the weaker flovirs. The 
exception noted was flour 1008. which showed the largest volume in 
the baking test, had a greater initial sugar content than any of the 
other flours, but did nt)t produce as much dextrose as flours 1001 and 
1009 when digested with malt flour. Flour 1007. a clear flour of very 
poor baking qualities, milled from Canadian wheat stood fourth in the 
series in regard to initial sugar content. Under the action of the dias- 
tase in malt flour, however, the reducing >ugars increased out of all 
proportion to its baking strength and on the addition of .5000 grams 
of malt flour, it contained more reducing sugars than any of the other 
flours with a like concentration of malt flour. 

In general the initial >ugar content (digestion 1 hour without added 
diastases I indicated the baking qualities of the flour quite accurately. 
This in turn depends to a large extent uj^on the diastatic enzymes con- 
tained in the flour itself. From the data presented, it would seem that 
the starch of the strong flours was generally more easily converted 
than that of weak flours. This was not in\aria1)le. a> flour 1008. a [)ar- 
ticularly strong patent flour showed only a very slight increase in sol- 
uble sugars when dige>ted with apjiroximately 5 ])ercent malt flour, 
whereas fl<jur 1007. a clear flour of notably poor baking cpialities. 
showed a phen<imenal increase under the same cx])erimental condi- 
tions. 

The Production of Reducing Sugars in the Panary Fermentation of 
Bread and the Effects of Diastases Added in the Dough. 

In this phase of the investigation, flours 1008 a strong ])atent. 1003 a 
weak flour, and 1002 a clear flour of good baking strength, were used. 
The data in Tables Xl\' to XXI and in Figures 6. 7 and 8 show how 
these three typical flours behave with regard to producing reducing 
sugars, when fermented normally and with dififerent amounts of added 
malt preparations. In order to have a check on the reducing sugars 
actually produced during the fermentation period, a check series was 
run at the same time, identical in all respects but having no yeast. In 
every instance where a diastatic enzyme w'as added, there was an 
increase in reducing sugars, over that of the normal dough, throughout 
the fermentation period. 

Flour 1008 had by far the greater amounts of reducing sugars, when 
the same additions of malt flour or malt extract were made, than did 

56 



the other two flours. Flour 1002 was next and 1003 wa^ at the bot- 
tom of the list. The curves in Figure 6, representing the production 
of sugars in Flour 1008, are all of the same general type, that is, the 
yeast dough curves are alike and the "no yeast' curves are alike. In 
the yeast doughs the peak of sugar formation or the point where the 
diastases were producing as much available sugar as the yeast was us- 
ing up, was at the first punch after two and one-half hours of fermen- 
tation. From this time on the }east seems to be stimulated and uses 
up the sugar faster than it is produced, steadily diminishing the sur- 
plus that the diastases have piled up in the first half of the fermenta- 
tion period. In the doughs which have no yeast, conversion seems to 
be slightly faster in the later stages of fermentation than in the begin- 
ning. If this is the case in the yeast dough, the yeast undoubtedly 
increases in activity as fermentation proceeds. \Mien 3 ])ercent malt 
extract is used, the sugar content is decidedly higher than in any of 
the other doughs. This is of decided advantage as the yeast has a 
large surplus of sugars to draw from. 

The curves for flour 1002 (Figure 8), are \ery similar to those for 
flour 1008 (Figure 6), with the exception that sugar production ap- 
peared to have reached a maximum at the end of 1 hour of fermenta- 
tion in the dough to which no diastattc enzymes were added and the 
one to which 3 percent malt extract was added. The doughs, to which 
1.5 and 4.0 percent malt flour were added, reached their maximum of 
sugar production at the first punch after a fermentation period of 2 
hours and fifteen minutes. The amounts of sugars produced by the 
diastatic enzymes are remarkably constant during the time of fermen- 
tation, with the greatest conversion produced l)y the addition of 4 per- 
cent malt flour. As in the preceding series, the dough to which the 
malt extract was added showed a larger amctunt of reducing sugars 
throughout the entire fermentation period. 

A difference in the shape of the cur\es is at once noticed (Figure 7) 
where the sugars produced, in the fermentation of flour 1003, are fol- 
lowed. Instead of an initial increase in reducing sugars, as w\as the 
case with the preceding strong flours, the yeast utiltizes all available 
sugars immediately, with a continued decrease in sugar content as fer- 
mentation proceeded. A slight increase in reducing sugars was se- 
cured by the use of 4 percent malt flour, which reached the highest 
point after 2 hours fermentation. The addition of 3 percent malt 
extract seemed to be able to convert enough starch to hold the avail- 
able reducing sugars constant for one hour, but after this time the 
yeast increased in activity and the available sugars dropped off rap- 
idly. 

.57 



From an iiibpectiun of Tables X1\'-XV1 and XXl-XXi\', and Fig- 
ures 6 and 8, it appeared that an addition of diastatic enzymes to a 
dough resulted in a surplus of reducing sugars during the earlier 
stages of fermentatic^n. This surplus was used up in the later stages 
of fermentation along with the sugars simultaneously produced by the 
diastatic enzymes. It also appeared that yeast activity was increased 
to a considerable extent when the carbon dioxide was punched out 
of the dough ; at least it seems to have been coincident with the punch- 
ing of the dough in the case of the stronger flours. It was also evi- 
dent from the auKnints of reducing sugars available, that the flours 
which showed good l)aking qualities had a greater diastatic activity 
than did the flour of poor baking strength. The malt preparations 
when added to weak flours produced less sugars in proportion than 
when added to strong flours. Whether or not the starch of the former 
was harder to hydrolyze than that of the latter is problematical but 
the data presented in section 2, and also in this section, might be taken 
as indicating such a possibility. 
Effects of the Proteolytic Enzymes, Contained in Malt Preparations, 

Upon the Viscosity of Strong and Weak Flours Following the 

Addition of Various Amounts of N 1 Lactic Acid. 

A very decided difference in the \ isco>ity of a flour- water suspension 
was noted when a flour was digested, with and without malt flour, for 
different periods of time, as sliown in Tal)le XXV, and illustrated 
graphically in Figure 9. Tlu' higher the concentration of malt flour 
added, the lower was the resulting viscosity reading, and when diges- 
tion was carried out for varied lengths of lime, a .steady decrease in 
viscosity occurred as time of digestion progressed. This was very 
noticeable as the percentage of malt flour was increased. 

When flours of different baking strengths were digested with in- 
creasing amounts of malt flour and malt extract, the viscosity of their 
suspensions in water (])lus lactic acid) decreased (juite decidedly as 
shown in Tables XXVI and XX\'1I, and graphically in Figures 10 and 
lOA. The strongest flours groui)ed themselves, and their suspen- 
.sions in acidified water ha\e a much liigher viscosity tlian those of 
the medium or weak flours, and when treated w ith 4 percent of malt 
flour or malt extract the strength of the flours was indicated by its 
position on the curve. The malt extract used did not decrease the viscosity 
as much as a like concentration of malt flour, and the conclusion was 
that it did not contain as large an amount of proteolytic enzymes as 
did the malt flour. It might be expected that the stronger flours 
would not show as great a decrease in viscosity as the weak flours. 
When digested with 4 jier cent malt flour or malt extract over the 
range given in Tables XX\T and XX\TI, but tlie oi)posite seems to be 

58 



actually the case. Flour 1008, the strongest flour in the series, showed 
a decrease in viscosity of 49° MacMichael, when digested with 4 per- 
cent ftxtract, while the decrease found for fiour 1003 under the same 
conditions is 22° and 24° M. respectively. Clear flour 1007 is interme- 
diate in tliis particular and showed a decrease of 36° MacMichael, 
when digested with 4 percent malt flour. 

Although it has been demonstrated that salts have a profound in- 
fluence upon the viscosity of flour- water suspensions, the results in 
Table XXYIII show that while the viscosity readings were very much 
higher in a flour-water suspension, from which the salts have been 
washed out, the same relative values hold, and the results recorded 
above were not vitiated by the electrolyte content of the flours. This 
has been demonstrated in another way where a flour was digested 
alone for four hours, with 4 percent malt flour for four hours, and an- 
other sample digested alone for three and one-half hours and at the end 
of this time 4 percent malt flour was added and digested thirty min- 
utes longer. It was thought that the salts of the added malt flour 
w(nild be extracted in thirty minutes and would exert their maximum 
effect in depressing the viscosity. Also, that in this length of time 
only a small amount of proteolytic activity would take place, thus 
showing a difference in viscosity between the flour which was digested 
four hours with 4 percent malt flour and the other which was digested 
three and one-half hours alone, and thirty minutes with 4 percent malt 
flour. These expectations were justitied, as demonstrated in Table 
XXIX, where the flour digested alone gaves a reading of 145°M., and 
that digested wath 4 i)ercent malt flour for four hours gave a reading 
of 81 °M., while that digested alone for three and one half hours and 
then thirty minutes more with malt flour gave a reading of 127°M. 
These data show that the increase in viscosity was not due entirely to 
the electrolytes but to the partial disintegration of the protein 

From the data presented in Tables XXVI and XXVH it has been 
shown that suspensions of strong flours in water have a higher viscos- 
ity than weak flours when digested with and without added malt prep- 
arations. It has also been shown that suspensions of strong flours in 
water show a greater decrease in viscosity than do similar suspensions 
of weak flours when digested with malt preparations and that the de- 
creases in viscosity recorded al)ove were not due entirely to the elec- 
trolyte content of the flours but to the cleavage of the gluten, thus de- 
creasing its imbibitional capacity and consecpiently its viscosity. 
The Gas Producing Capacities of Strong and Weak Flours and the 
Effect of Added Malt Extract Upon Them. 

Wood has shown that the gas produced by a flour, especially in the 
later stages of fermentation, was a factor in strength, while Baker and 

59 



llulton have shown that in some cases a weak flour produces as much, 
and in some cases even more, gas than does a strong flour. They be- 
lieved that weak flours were deficient in liquifying enzymes and that 
an addition of liquifying enzymes would increase the gas production 
of a weak flour to a considerable extent, while they would have little 
or no efifect upon a strcmg flour. The data in Tal)le XXX supports 
their theorv and shtnvs that flours 1008 and 1009, which showed very 
g(K)d baking qualities, did not increase to any extent in gas producing 
capacity when malt extract was added, while flour 1002, a strong clear 
fl(nir increased only 9 cc. under tlie same conilitions, and 1007, a clear 
flour of poor baking quality increased 37 cc. under the same treatment. 
l"he test seems to be conclusi\e l)y the increase shown by flour 1003, 
a notably weak flour, ^\•hich increased 80 cc. when 1 percent malt ex- 
tract was added. 

The Changes in Hydrogen Ion Concentration Taking Place During 
the Fermentation of the Dough. 

The clianges in h}-drogen ion concentration taking place during the 
fermentation of the dough, are recorded in Table XXXI and show that 
steady increase in hydorgen ion concentration takes ])lace as fermen- 
tation proceeds. With two excei)tions the doughs wlien ready ("or the 
oven had a hydrt)gen ion concentration of ap])roximately pll 5. The 
two flours which had a higher ])I1 were cle;ir flours ot \ cry jxxir 
baking strength. 

The Effects of Malt Flour and Malt Extract Upon the Baking Value 

of Flour. 

In flour 1001, a strong patent flour, the xolmne was considerably 
increased by the use of 2.5 i)ercent malt flour. This advantage was 
materially offset by the decrease in color. A\"ith the use of the malt ex- 
tract, the volume increased with additions u]) ti> 3 i)ercent with not 
much decrease in color, while a sweet honey-like flavor is evident 
which adds to the value of the loaf. The data in 1\Hble XXXIII 
shows that the baking qualities of the flour were improved when 3 per- 
cent malt extract was used. 

Flour 1002, a fairlv strong clear flour, increased in volume with the 
addition of malt flour. The greatest volume was secured by the use 
of 2 percent malt flour for the weight of the dough baked out. While 
the grain and texture were uniform throughout, the decrease in color 
value ofYset the advantages secured by the increase in volume. In 
using malt extract the greatest volume was secured by the use of 1.5 
and 2.0 percent, and decidedly the best loaves were thus produced, 
since texture and grain increased in fineness as the amount of malt ex- 
tract increased. The slight decrease in color value was not a serious 

60 



objection and the addition of 1.5 to 2.0 percent malt extract had a 
decided beneficial effect upon the baking qualities of flour 1002 

With the use of 1.5 percent malt flour the largest volume was se- 
cured in baking flour 1003. As the grain was coarse and the color off, 
however, the advantages gained by the increase in volume were off- 
set. A decided increase in volume and grain was secured by the use 
of 1.5 percent malt extract in this weak flour in my opinion, the baking 
quality of this flour was thus greatly increased. 

In clear flour 1007, the use of 1.5 percent malt flour increased the 
volume as well as the texture and grain, and in this flour the addition 
of malt flour was beneficial. The use of 3 percent malt extract gave a 
decided improvement in the baking qualities of flmir 1007 as far as 
volume, grain, texture and color is concerned. 

In flour 1008, the strongest flour of the series, the use of 1.5 percent 
malt flour improved the texture and grain but darkened the color con- 
siderably. The use of malt flour did not increase the baking qualities 
of this flour, while on the other hand the use of 3 percent malt extract 
increased the volume slightly, improved the texture and grain thus 
improving the baking qualities to a marked extent. 



61 



IV. SUMMARY 

This paper deals witli the effects of diastatic ferments upon the 
strength of wheat flours. Tables and graphs have been presented, 
showing: 

1. Optimum tem,perature fur the diastatic actixity of the malt flour 
used was at temperature oi 65°C. 

2. Optimum hydrogen ion concentration for tlie diastatic enzymes 
in malt tlour was at a pH of 4.2C). 

3. Constant diastatic acti\it\' was shown by the malt flour over a 
perio(l of eight hour> when digeNted at 27^C 

4. Concentrations of added diastase exert their greatest effect in 
tlie first 10 percent of added malt Hour, giving an increase in dextrose 
from 1.63 to 3.<y percent. 

5. Diastatic ferments when added to wheat Hours increase the 
reducing sugar>. when dige^ted at 27' C for 1 hour. The strong 
flours showed a higiier sugar content and greater diastatic activity 
than did the weaker flours, ddie starch of the strong flours appeared 
to be more ea>il}' hydroly/ed by diastatic ferments than that of the 
weaker flours. 

6. Addition of (lia>tatic ferments to a dough con\ert tlie starch 
to reducing sugars and in the earlier stages of fermentation, produce 
a surj^lus of fermentable sugars in the doughs made from strong 
flours. This surplu> :^oon di>a|)p(.-ar> as the acti\ity oi the yeast 
increases, and at the end of the ft-rmentatiou ])eriod the dough is 
nearly depleted of a\ailal)le sugars. 

7. Susi)ension-^ of strong flours in water had a higher \iscosity (on 
the addition of lactic acid i than simibir >uspen>ions of weak flours 
when incubated alone or with added <liastatic ferments in the form 
(.)f lualt flour and malt extract. The course of ])roteo|ytic activity 
could be accurateh- followed by the cliange in viscosity when wheat 
flour was digested with added malt flour. The ])resence of naturall_\' 
occurring salts of the wheat flour-- did not \itiate the \iscosity readings. 

8. Gas producing capacit\- of weak flours was greatly increased 
when fermented with added malt extract. This was not the case 
wdien strong flours were fermented with a<lded malt extract. 

9. Hydrogen ion concentration of the dough steadily increased as 
fermentation i)roceeded. With two exceptions, the doughs were at 
api^roximately a pll of 5.0 when ready for the oven. 

10. Addition of malt flour and malt extract to doughs increased 
the volume of the residting bread. In all cases the use of malt extract 
gave a superior loaf of l)read in xolume, grain and texture, thus 
increasing the baking strength of the flours. 

62 



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65 



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66 



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OH 



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72 



BIOGRAPHICAL. 

Ferdinand Albert Collatz was born in Duluth, Minnesota. He 
graduated from the Duluth Central Hig-h School in June, 1914, and 
entered the University of Minnesota the same fall, where he received 
the degree of Bachelor of Science in June, 1918. Shortly after this 
he entered the Army and was assigned to the Piiysiological Labora- 
tory at the Lakeside Hospital, Cleveland, Ohio, under the direction 
of Major Roy G. Pierce. During 1919-1920, he held the position of 
Assistant in Agricultural Biochemistry. University of Minnesota, 
and in June. 1920, received the degree of Master of Science from this 
department. During 1920-21 he held the American Institute of Bak- 
ing Research Fellowship, where the experimental work in this Thesis 
was done, at the same time continuing his graduate work in the de- 
partment of Agricultural Biochemistry, University of Minnesota. 
Here he studied for the degree of Doctor of Philosophy. 

Major subject, Biochemistry. 

Minor subject. Botany. 

Member of Sigma Xi, Phi Lambda Upsilon, Gamma Sigma Delta. 
Gamma Alpha; Mcmbt-r of llie American Chemical .Society. 



73 



ACKNOWLEDGMENT. 

This imotigaliim was carried out under \hv direclioii of Dr. Ro.ss 
.■\iken (lortner. 'Jdic author lakes ilii> opportuuitx- to ex])ress his 
ai)i)reciation and gratitude for the help and encourajj;"enu-nt wliich wa-- 
so i^ladly g'iven durin<;' the lime this work was in progress. 

F. .\. COLL.ATZ. 



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